Seed plants characterized by delayed seed dispersal

The present invention provides a non-naturally occurring seed plant that is characterized by delayed seed dispersal due to ectopic expression of a nucleic acid molecule encoding an AGL8-like gene product. Further provided herein is a non-naturally occurring seed plant, such as an agl1 agl5 double mutant, that is characterized by delayed seed dispersal due to suppression of AGL1 and AGL5 expression in the seed plant. The invention also provides a substantially purified dehiscence zone-selective regulatory element, which includes a nucleotide sequence that confers selective expression upon an operatively linked nucleic acid molecule in the valve margin or dehiscence zone of a seed plant. Also provided by the invention are kits for producing a transgenic seed plant characterized by delayed seed dispersal, such kits containing a dehiscence zone-selective regulatory element.

BACKGROUND OF THE INVENTION
 1. FIELD OF THE INVENTION
 The present invention relates generally to plant molecular biology and
 genetic engineering and more specifically to the production of genetically
 modified seed plants in which the natural process of dehiscence is
 delayed.
 2. BACKGROUND INFORMATION
 Rapeseed is one of the most important oilseed crops after soybeans and
 cottonseed, representing 10% of the world oilseed production in 1990.
 Rapeseed contains 40% oil, which is pressed from the seed, leaving a
 high-protein seed meal of value for animal feed and nitrogen fertilizer.
 Rapeseed oil, also known as canola oil, is a valuable product,
 representing the fourth most commonly traded vegetable oil in the world.
 The production of oilseeds, meal and oil from rapeseed plants has been
 increasing continuously for the last 30 years for food and feed grains,
 mainly by expansion of the area under cultivation. Most northern European
 countries produce rapeseed as their main edible oil crop. By the year
 2000, China is expected to be the leading producer with 9.2 metric tons
 (Mt; 26%); followed by India with 7.8 Mt (22%); the European Community (12
 countries), with 7.6 Mt (21%); Canada, 3.8 Mt (11%) and eastern Europe
 with 2.6 Mt (7%).
 Unfortunately, the yield of seed from rapeseed and related plants is
 limited by pod dehiscence, which is a process that occurs late in fruit
 development whereby the pod is opened and the enclosed seeds released.
 Degradation and separation of cell walls along a discrete layer of cells
 dividing the two halves of the pod, termed the "dehiscence zone," result
 in separation of the two halves of the pod and release of the contained
 seeds. Seed "shattering," whereby seeds are prematurely shed through
 dehiscence before the crop can be harvested, is a significant problem
 faced by commercial seed producers and represents a loss of income to the
 industry. Adverse weather conditions can exacerbate the process of
 dehiscence, resulting in greater than 50% loss of seed yield.
 Attempts to solve this problem over the past 20 years have focused on the
 breeding of shatter-resistant varieties. However, these plant hybrids are
 frequently sterile and lose favorable characteristics that must be
 regained by backcrossing, which is both time-consuming and laborious.
 Other strategies to alleviate pod shattering include the use of chemicals
 such as pod sealants or mechanical techniques such as swathing to reduce
 wind-stimulated shattering. To date, however, a simple method for
 producing genetically modified seed plants that do not open and release
 their seeds prematurely has not been described.
 Thus, a need exists for identifying genes that regulate the dehiscence
 process and for developing genetically modified seed plant varieties in
 which the natural seed dispersal process is delayed. The present invention
 satisfies this need and provides related advantages as well.
 SUMMARY OF THE INVENTION
 The present invention provides a non-naturally occurring seed plant that is
 characterized by delayed seed dispersal due to ectopic expression of a
 nucleic acid molecule encoding an AGL8-like gene product. The AGL8-like
 gene product can have, for example, substantially the amino acid sequence
 of an AGL8 ortholog such as Arabidopsis AGL8 (SEQ ID NO:2). Particularly
 useful seed plants of the invention, which are characterized by delayed
 seed dispersal, include members of the Brassicaceae, such as rapeseed, and
 members of the Fabaceae, such as soybeans, peas, lentils and beans.
 In one embodiment, the invention provides a transgenic seed plant that is
 characterized by delayed seed dispersal due to ectopic expression of a
 nucleic acid molecule encoding an AGL8-like gene product. In a transgenic
 seed plant of the invention, the nucleic acid molecule encoding the
 AGL8-like gene product can be operatively linked to an exogenous
 regulatory element. Useful exogenous regulatory elements include
 constitutive regulatory elements and dehiscence zone-selective regulatory
 elements. In particular, the exogenous regulatory element can be a
 dehiscence zone-selective regulatory element that is an AGL1 regulatory
 element or an AGL5 regulatory element.
 In another embodiment, the invention provides a non-naturally occurring
 seed plant that is characterized by delayed seed dispersal due to
 suppression of both AGL1 and AGL5 expression in the seed plant. Such a
 non-naturally occurring seed plant characterized by delayed seed dispersal
 can be, for example, an agl1 agl5 double mutant.
 The present invention further provides a tissue derived from a
 non-naturally occurring seed plant of the invention. In one embodiment,
 the invention provides a tissue derived from a non-naturally occurring
 seed plant that has an ectopically expressed nucleic acid molecule
 encoding an AGL8-like gene product and is characterized by delayed seed
 dispersal. In another embodiment, the invention provides a tissue derived
 from a non-naturally occurring seed plant in which AGL1 expression and
 AGL5 expression each are suppressed, where the seed plant is characterized
 by delayed seed dispersal.
 Methods of producing a non-naturally occurring seed plant characterized by
 delayed seed dispersal also are provided herein. Such methods entail
 ectopically expressing a nucleic acid molecule encoding an AGL8-like gene
 product in the seed plant, whereby seed dispersal is delayed due to
 ectopic expression of the nucleic acid molecule.
 The invention also provides a substantially purified dehiscence
 zone-selective regulatory element, comprising a nucleotide sequence that
 confers selective expression upon an operatively linked nucleic acid
 molecule in the valve margin or dehiscence zone of a seed plant, provided
 that the dehiscence zone-selective regulatory element does not have a
 nucleotide sequence consisting of nucleotides 1889 to 2703 of SEQ ID NO:4.
 The dehiscence zone-selective regulatory element can be, for example, an
 AGL1 regulatory element or AGL5 regulatory element.
 Further provided is a plant expression vector containing a dehiscence
 zone-selective regulatory element that confers selective expression upon
 an operatively linked nucleic acid molecule in the valve margin or
 dehiscence zone of a seed plant, provided that the dehiscence
 zone-selective regulatory element does not have a nucleotide sequence
 consisting of nucleotides 1889 to 2703 of SEQ ID NO:4. If desired, a plant
 expression vector can contain a nucleic acid molecule encoding an
 AGL8-like gene product in addition to the dehiscence zone-selective
 regulatory element.
 The invention also provides a kit for producing a transgenic seed plant
 characterized by delayed seed dispersal, such kit containing a dehiscence
 zone-selective regulatory element that confers selective expression upon
 an operatively linked nucleic acid molecule in the valve margin or
 dehiscence zone of a seed plant, provided that said dehiscence
 zone-selective regulatory element does not have a nucleotide sequence
 consisting of nucleotides 1889 to 2703 of SEQ ID NO:4. In a kit of the
 invention, the dehiscence zone-selective regulatory element can be, if
 desired, operatively linked to a nucleic acid molecule encoding an
 AGL8-like gene product.

DETAILED DESCRIPTION OF THE INVENTION
 The present invention provides a non-naturally occurring seed plant that is
 characterized by delayed seed dispersal due to ectopic expression of a
 nucleic acid molecule encoding an AGL8-like gene product. The AGL8-like
 gene product can have, for example, substantially the amino acid sequence
 of an AGL8 ortholog such as Arabidopsis AGL8 (SEQ ID NO:2).
 The fruit, a complex structure unique to flowering plants, mediates the
 maturation and dispersal of seeds. In most flowering plants, the fruit
 consists of the pericarp, which is derived from the ovary wall, and the
 seeds, which develop from fertilized ovules. Arabidopsis, which is typical
 of the more than 3000 species of the Brassicaceae, produces fruit in which
 the two carpel valves (ovary walls) are joined to the replum, a visible
 suture that divides the two carpels. The structure of an Arabidopsis
 gynoecium around the time of pollination, including the carpel valves and
 replum, is shown in FIG. 1.
 Pod dehiscence or shatter occurs late in fruit development in a wide
 spectrum of important plant crops such as oilseed rape (Brassica napus L.)
 and is a process of economic importance that can lead to significant
 losses in seed yield. In oilseed rape, dehiscence involves the breakdown
 of cell wall material in a discrete cell layer known as the "dehiscence
 zone," which is a region of only one to three cells in width that extends
 along the entire length of the valve/replum boundary (Meakin and Roberts,
 J. Exp. Botany 41:995-1002 (1990)). As the cells in the dehiscence zone
 separate from one another, the valves detach from the replum, allowing
 seeds to be dispersed (see FIG. 2).
 The plant hormone ethylene is produced by developing seeds and appears to
 be an important regulator of the dehiscence process. One line of evidence
 supporting a role for ethylene in regulation of dehiscence comes from
 studies of fruit ripening, which, like fruit dehiscence, is a process
 involving the breakdown of cell wall material. In fruit ripening, ethylene
 acts in part by activating cell wall degrading enzymes such as
 polygalacturonase (Theologis et al., Develop. Genetics 14:282-295 (1993)).
 Moreover, in genetically modified tomato plants in which the ethylene
 response is blocked, such as transgenic tomato plants expressing antisense
 polygalacturonase, there is a significant delay in fruit ripening (Lanahan
 et al., The Plant Cell 6:521-530 (1994); Smith et al., Nature 334:724-726
 (1988)).
 In dehiscence, ultrastructural changes that culminate in degradation of the
 middle lamella of dehiscence zone cell walls weaken rapeseed pods and
 eventually lead to pod shatter. As in fruit ripening, hydrolytic enzymes
 including polygalacturonases play a role in this programmed breakdown. For
 example, in oilseed rape, a specific endo-polygalacturonase, RDPG1, is
 upregulated and expressed exclusively in the dehiscence zone late in pod
 development (Petersen et al., Plant Mol. Biol. 31:517-527 (1996), which is
 incorporated herein by reference). Ethylene may regulate the activity of
 hydrolytic enzymes involved in the process of dehiscence as it does in
 fruit ripening (Meakin and Roberts, J. Exp. Botany 41:1003-1011 (1990),
 which is incorporated herein by reference). Yet, until now, the proteins
 that control the process of dehiscence, such as those regulating the
 relevant hydrolytic enzymes, have eluded identification.
 The present invention is directed to the surprising discovery that the AGL8
 transcription factor regulates the process of dehiscence. As disclosed
 herein, Arabidopsis plants were transformed with an AGL8 cDNA under
 control of a 35S cauliflower mosaic virus (CaMV) constitutive promoter
 such that AGL8 was ectopically expressed throughout the transformed plant.
 In particular, AGL8, which is normally expressed in the carpel valves, was
 ectopically expressed in the replum, which is a small strip of cells
 separating the two valves in a mature fruit. As a consequence of such
 ectopic expression, the replum of the fruit was absent, with the cells of
 the outer replum replaced by cells having characteristics of valve
 identity, demonstrating that, in this context, AGL8 expression is
 sufficient to specify valve cell fate. Furthermore, ectopic expression of
 the AGL8 cDNA produced a transgenic plant in which the dehiscence zone
 failed to develop normally, resulting in delayed seed dispersal (see
 Example I). Whereas wild type Arabidopsis produced fruit that opened and
 released seeds on or about 14 days after pollination, transformed
 Arabidopsis ectopically expressing AGL8 produced fruit in which seed
 dispersal was postponed, or in which the seeds were never released unless
 the fruit was opened manually (see FIG. 3). Thus, for the first time, seed
 plants were genetically modified to delay the natural process of
 dehiscence.
 The present invention also relates to the surprising discovery that an agl1
 agl5 double mutant seed plant has a delayed seed dispersal phenotype that
 is strikingly similar to the AGL8 gain-of-function phenotype. As disclosed
 herein, loss-of-function mutations in the AGL1 and AGL5 genes were
 produced by disruptive T-DNA insertion and homologous recombination (see
 Example II). In the resulting agl1 agl5 double mutant plants, the
 dehiscence zone failed to develop normally, and the mature fruits did not
 undergo dehiscence (see FIG. 5). Thus, AGL1 or AGL5 gene expression is
 required for development of the dehiscence zone. These results indicate
 that AGL1, AGL5 and AGL8 regulate pod dehiscence and that manipulation of
 AGL1, AGL5 and AGL8 expression can allow the process of pod shatter to be
 controlled.
 Thus, the present invention provides a non-naturally occurring seed plant
 that is characterized by delayed seed dispersal due to ectopic expression
 of a nucleic acid molecule encoding an AGL8-like gene product. The
 AGL8-like gene product can have, for example, substantially the amino acid
 sequence of an AGL8 ortholog such Arabidopsis AGL8 (SEQ ID NO:2).
 As used herein, the term "non-naturally occurring," when used in reference
 to a seed plant, means a seed plant that has been genetically modified by
 man. A transgenic seed plant of the invention, for example, is a
 non-naturally occurring seed plant that contains an exogenous nucleic acid
 molecule encoding an AGL8-like gene product and, therefore, has been
 genetically modified by man. In addition, a seed plant that contains, for
 example, a mutation in an endogenous AGL8-like gene product regulatory
 element or coding sequence as a result of calculated exposure to a
 mutagenic agent, such as a chemical mutagen, or an "insertional mutagen,"
 such as a transposon, also is considered a non-naturally occurring seed
 plant, since it as been genetically modified by man. In contrast, a seed
 plant containing only spontaneous or naturally occurring mutations is not
 a "non-naturally occurring seed plant" as defined herein and, therefore,
 is not encompassed within the invention. One skilled in the art
 understands that, while a non-naturally occurring seed plant typically has
 a nucleotide sequence that is altered as compared to a naturally occurring
 seed plant, a non-naturally occurring seed plant also can be genetically
 modified by man without altering its nucleotide sequence, for example, by
 modifying its methylation pattern.
 The term "ectopically," as used herein in reference to expression of a
 nucleic acid molecule encoding an AGL8-like gene product, refers to an
 expression pattern that is distinct from the expression pattern in a wild
 type seed plant. Thus, one skilled in the art understands that ectopic
 expression of a nucleic acid encoding an AGL8-like gene product can refer
 to expression in a cell type other than a cell type in which the nucleic
 acid molecule normally is expressed, or at a time other than a time at
 which the nucleic acid molecule normally is expressed, or at a level other
 than the level at which the nucleic acid molecule normally is expressed.
 In wild type Arabidopsis, for example, AGL8 expression is normally
 restricted during the later stages of floral development to the carpel
 valves and is not seen in the replum, which is the small strip of cells
 separating the carpel valves. However, under control of a constitutive
 promoter such as the cauliflower mosaic virus 35S promoter, AGL8 is
 expressed in the replum and, additionally, is expressed at higher than
 normal levels in other tissues such as valve margin and, thus, is
 ectopically expressed.
 The term "delayed," as used herein in reference to the timing of seed
 dispersal in a fruit produced by a non-naturally occurring seed plant of
 the invention, means a significantly later time of seed dispersal as
 compared to the time seeds normally are dispersed from a corresponding
 seed plant lacking an ectopically expressed nucleic acid molecule encoding
 an AGL8-like gene product. Thus, the term "delayed" is used broadly to
 encompass both seed dispersal that is significantly postponed as compared
 to the seed dispersal in a corresponding seed plant, and to seed dispersal
 that is completely precluded, such that fruits never release their seeds
 unless there is human or other intervention.
 It is recognized that there can be natural variation of the time of seed
 dispersal within a seed plant species or variety. However, a "delay" in
 the time of seed dispersal in a non-naturally occurring seed plant of the
 invention readily can be identified by sampling a population of the
 non-naturally occurring seed plants and determining that the normal
 distribution of seed dispersal times is significantly later, on average,
 than the normal distribution of seed dispersal times in a population of
 the corresponding seed plant species or variety that does not contain an
 ectopically expressed nucleic acid molecule encoding an AGL8-like gene
 product. Thus, production of non-naturally occurring seed plants of the
 invention provides a means to skew the normal distribution of the time of
 seed dispersal from pollination, such that seeds are dispersed, on
 average, at least about 1%, 2%, 5%, 10%, 30%, 50% or 100% later than in
 the corresponding seed plant species that does not contain an ectopically
 expressed nucleic acid molecule encoding an AGL8-like gene product.
 A delay in seed dispersal of even one to two days can be valuable in
 increasing the amount of seed successfully harvested from a seed plant. In
 canola rapeseed, for example, dehiscence normally occurs about 8 weeks
 post-pollination. In a non-naturally occurring canola rapeseed that
 ectopically expresses an AGL8-like gene product, dehiscence can occur one
 to two days later than in the wild type variety, allowing a significantly
 greater percentage of the seed crop to be harvested rather than lost
 through uncontrolled seed dispersal.
 The present invention relates to the use of nucleic acid molecules encoding
 particular "AGAMOUS-LIKE" or "AGL" gene products. AGAMOUS (AG) is a floral
 organ identity gene, one of a related family of transcription factors
 that, in various combinations, specify the identity of the floral organs:
 the petals, sepals, stamens and carpels (Bowman et al., Devel. 112:1-20
 (1991); Weigel and Meyerowitz, Cell 78:203-209 (1994); Yanofsky, Annual
 Rev. Plant Physiol. Mol. Biol. 46:167-188 (1995)). The AGAMOUS gene
 product is essential for specification of carpel and stamen identity
 (Bowman et al., The Plant Cell 1:37-52 (1989); Yanofsky et al., Nature
 346:35-39 (1990)). Related genes have recently been identified and denoted
 "AGAMOUS-LIKE" or "AGL" genes (Ma et al., Genes Devel. 5:484-495 (1991);
 Mandel and Yanofsky, The Plant Cell 7:1763-1771 (1995), which is
 incorporated herein by reference).
 AGL8, like AGAMOUS and other AGL genes, is characterized, in part, in that
 it is a plant MADS box gene. The plant MADS box genes generally encode
 proteins of about 260 amino acids including a highly conserved MADS domain
 of about 56 amino acids (Riechmann and Meyerowitz, Biol. Chem.
 378:1079-1101 (1997), which is incorporated herein by reference). The MADS
 domain, which was first identified in the Arabidopsis AGAMOUS and
 Antirrhimum majus DEFICIENS genes, is conserved among transcription
 factors found in humans (serum response factor; SRF) and yeast (MCM1;
 Norman et al., Cell 55:989-1003 (1988); Passmore et al., J. Mol. Biol.
 204:593-606 (1988), and is the most highly conserved region of the MADS
 domain proteins. The MADS domain is the major determinant of sequence
 specific DNA-binding activity and can also perform dimerization and other
 accessory functions (Huang et al., The Plant Cell 8:81-94 (1996)). The
 MADS domain frequently resides at the N-terminus, although some proteins
 contain additional residues N-terminal to the MADS domain.
 The "intervening domain" or "I-domain," located immediately C-terminal to
 the MADS domain, is a weakly conserved domain having a variable length of
 approximately 30 amino acids (Purugganan et al., Genetics 140:345-356
 (1995)). In some proteins, the I-domain plays a role in the formation of
 DNA-binding dimers. A third domain present in plant MADS domain proteins
 is a moderately conserved 70 amino acid region denoted the "keratin-like
 domain" or "K-domain." Named for its similarity to regions of the keratin
 molecule, the structure of the K-domain appears capable of forming
 amphipathic helices and may mediate protein-protein interactions (Ma et
 al., Genes Devel. 5:484-495 (1991)). The most variable domain, both in
 sequence and in length, is the carboxy-terminal or "C-domain" of the MADS
 domain proteins. Dispensable for DNA binding and protein dimerization in
 some MADS domain proteins, the function of this C-domain remains unknown.
 Arabidopsis AGL8 is a 242 amino acid MADS box protein (see FIG. 6; SEQ ID
 NO:2; Mandel and Yanofsky, supra, 1995). The AGL8 MADS domain resides at
 amino acids 2 to 56 of SEQ ID NO:2. The K-domain of AGL8 resides at amino
 acids 92 to 158 of SEQ ID NO:2.
 In wild-type Arabidopsis, AGL8 RNA accumulates in two distinct phases, the
 first occurring during inflorescence development in the stem and cauline
 leaves and the second in the later stages of flower development (Mandel
 and Yanofsky, supra, 1995). In particular, AGL8 RNA is first detected in
 the inflorescence meristem as soon as the plant switches from vegetative
 to reproductive development. As the inflorescence stem elongates, AGL8 RNA
 accumulates in the inflorescence meristem and in the stem. Secondly,
 although AGL8 is not detected in the initial stages (1 and 2) of flower
 development, AGL8 expression resumes at approximately stage 3 in the
 center of the floral dome in the region corresponding to the fourth
 (carpel) whorl. AGL8 expression is excluded from all other primordia and
 the pedicel. The time of AGL8 expression in the fourth carpel whorl
 generally corresponds to the time at which the organ identity genes
 APETALA3, PISTILLATA AND AGAMOUS begin to be expressed (Yanofsky et al.,
 Nature 346:35-39 (1990); Drews et al., Cell 65:991-1002 (1991); Jack et
 al., Cell 68:683-697 (1992); Goto and Meyerowitz, Genes Devel. 8:1548-1560
 (1994)). At later stages, AGL8 expression becomes localized to the carpel
 walls, in the region that constitutes the valves of the ovary, and is
 absent from nearly all other cell types of the carpel. No AGL8 RNA
 expression is detected in the ovules, stigmatic tissues or the septum that
 divides the ovary. Thus, in nature, AGL8 expression during the later
 stages of floral development is restricted to the valves of the carpels
 and to the cells within the style.
 As used herein, the term "AGL8-like gene product" means a gene product that
 has the same or similar function as Arabidopsis AGL8 such that, when
 ectopically expressed in a seed plant, the normal development of the
 dehiscence zone is altered, and seed dispersal is delayed. An AGL8-like
 gene product can have, for example, the ability to convert cells of the
 outer replum to a valve cell identity. Arabidopsis AGL8 (SEQ ID NO:2) is
 an example of an AGL8-like gene product as defined herein. As disclosed in
 Example I, ectopic expression of Arabidopsis AGL8 (SEQ ID NO:2) under
 control of a tandem CaMV 35S promoter, in which the intrinsic promoter
 element has been duplicated, alters formation of the dehiscence zone,
 thereby resulting in fruit characterized by a complete lack of seed
 dispersal. An AGL8-like gene product also can be characterized, in part,
 by its ability to interact with AGL1 and, additionally, its ability to
 interact with AGL5.
 An AGL8-like gene product generally is characterized, in part, by having an
 amino acid sequence that has at least about 50% amino acid identity with
 the amino acid sequence of Arabidopsis AGL8 (SEQ ID NO: 2). An AGL8-like
 gene product can have, for example, an amino acid sequence with greater
 than about 65% amino acid sequence identity with Arabidopsis AGL8 (SEQ ID
 NO:2), preferably greater than about 75% amino acid identity with
 Arabidopsis AGL8 (SEQ ID NO:2), more preferably greater than about 85%
 amino acid identity with Arabidopsis AGL8 (SEQ ID NO:2), and can be a
 sequence having greater than about 90%, 95% or 97% amino acid identity
 with Arabidopsis AGL8 (SEQ ID NO:2).
 Preferably, an AGL8-like gene product is orthologous to the seed plant
 species in which it is ectopically expressed. A nucleic acid molecule
 encoding Arabidopsis AGL8 (SEQ ID NO:2), for example, can be ectopically
 expressed in an Arabidopsis plant to produce a non-naturally occurring
 Arabidopsis variety characterized by delayed seed dispersal. Similarly, a
 nucleic acid molecule encoding canola AGL8 can be ectopically expressed in
 a canola plant to produce a non-naturally occurring canola variety
 characterized by delayed seed dispersal.
 A nucleic acid molecule encoding an AGL8-like gene product also can be
 ectopically expressed in a heterologous seed plant to produce a
 non-naturally occurring seed plant characterized by delayed seed
 dispersal. AGAMOUS-like gene products have been widely conserved
 throughout the plant kingdom; for example, AGAMOUS has been conserved in
 tomato (TAG1) and maize (ZAG1), indicating that orthologs of AGAMOUS-like
 genes are present in most, if not all, angiosperms (Pnueli et al., The
 Plant Cell 6:163-173 (1994); Schmidt et al., The Plant Cell 5:729-737
 (1993)). AGL8-like gene products such as AGL8 orthologs also can be
 conserved and can function across species boundaries to delay seed
 dispersal. Thus, ectopic expression of a nucleic acid molecule encoding
 Arabidopsis AGL8 (SEQ ID NO:2) in a heterologous seed plant within the
 Brassicaceae such as Brassica napus L. (rapeseed) or within the Fabaceae
 such as in Glycine (soybean) can alter normal development of the
 dehiscence zone, thereby resulting in delayed seed dispersal. Furthermore,
 a nucleic acid molecule encoding Arabidopsis AGL8 (SEQ ID NO:2), for
 example, can be ectopically expressed in more distantly related
 heterologous seed plants, including dehiscent seed plants as well as other
 dicotyledonous and monocotyledonous angiosperms and gymnosperms and, upon
 ectopic expression, can alter normal development of the dehiscence zone
 and delay seed dispersal in the heterologous seed plant.
 As used herein, the term "AGL8-like gene product" encompasses an active
 segment of an AGL8-like gene product, which is a polypeptide portion of an
 AGL8-like gene product that, when ectopically expressed, alters normal
 development of the dehiscence zone and delays seed dispersal. An active
 segment can be, for example, an amino terminal, internal or carboxy
 terminal fragment of Arabidopsis AGL8 (SEQ ID NO:2) that, when ectopically
 expressed in a seed plant, alters normal development of the dehiscence
 zone and delays seed dispersal. An active segment of an AGL8-like gene
 product can include, for example, the MADS domain and can have the ability
 to bind DNA specifically. The skilled artisan will recognize that a
 nucleic acid molecule encoding an active segment of an AGL8-like gene
 product can be useful in producing a seed plant of the invention
 characterized by delayed seed dispersal and in the related methods and
 kits of the invention described further below.
 An active segment of an AGL8-like gene product can be identified using the
 methods described in Example I or using other routine methodology.
 Briefly, a seed plant such as Arabidopsis can be transformed with a
 nucleic acid molecule under control of a constitutive regulatory element
 such as a tandem CaMV 35S promoter. Phenotypic analysis of the seed plant
 reveals whether a seed plant ectopically expressing a particular
 polypeptide portion is characterized by delayed seed dispersal. In
 transgenic plants in which seed dispersal is delayed, further analysis can
 be performed to confirm that normal development of the dehiscence zone has
 been altered. For analysis of a large number of polypeptide portions of an
 AGL8-like gene product, nucleic acid molecules encoding the polypeptide
 portions can be assayed in pools, and active pools subsequently subdivided
 to identify the active nucleic acid molecule.
 In one embodiment, the invention provides a non-naturally occurring seed
 plant that is characterized by delayed seed dispersal due to ectopic
 expression of a nucleic acid molecule encoding an AGL8-like gene product
 having substantially the amino acid sequence of an AGL8 ortholog. As used
 herein, the term "AGL8 ortholog" means an ortholog of Arabidopsis AGL8
 (SEQ ID NO:2) and refers to an AGL8-like gene product that, in a
 particular seed plant variety, has the highest percentage homology at the
 amino acid level to Arabidopsis AGL8 (SEQ ID NO:2). An AGL8 ortholog can
 be, for example, a Brassica AGL8 ortholog such as a Brassica napus L. AGL8
 ortholog, or a Fabacea AGL8 ortholog such as a soybean, pea, lentil, or
 bean AGL8 ortholog. An AGL8 ortholog from the long-day plant Sinapis alba,
 designated SaMADS B, has been described (Menzel et al., Plant J. 9:399-408
 (1996), which is incorporated herein by reference). Novel AGL8 ortholog
 cDNAs can be isolated from additional seed plant species using a
 nucleotide sequence as a probe and methods well known in the art of
 molecular biology (Glick and Thompson (eds.), Methods in Plant Molecular
 Biology and Biotechnology, Boca Raton, Fla.: CRC Press (1993); Sambrook et
 al. (eds.), Molecular Cloning: A Laboratory Manual (Second Edition),
 Plainview, N.Y.: Cold Spring Harbor Laboratory Press (1989), each of which
 is incorporated herein by reference).
 As used herein, the term "substantially the amino acid sequence," when used
 in reference to an AGL8 ortholog, is intended to mean a polypeptide or
 polypeptide segment having an identical amino acid sequence, or a
 polypeptide or polypeptide segment having a similar, non-identical
 sequence that is considered by those skilled in the art to be a
 functionally equivalent amino acid sequence. For example, an AGL8-like
 gene product having substantially the amino acid sequence of Arabidopsis
 AGL8 can have an amino acid sequence identical to the sequence of
 Arabidopsis AGL8 (SEQ ID NO:2) shown in FIG. 6, or a similar,
 non-identical sequence that is functionally equivalent. In particular, an
 amino acid sequence that is "substantially the amino acid sequence" of
 AGL8 can have one or more modifications such as amino acid additions,
 deletions or substitutions relative to the AGL8 amino acid sequence shown
 (SEQ ID NO:2), provided that the modified polypeptide retains
 substantially the ability to alter normal development of the dehiscence
 zone and delay seed dispersal when ectopically expressed in the seed
 plant. Comparison of sequences for substantial similarity can be performed
 between two sequences of any length and usually is performed with
 sequences between about 6 and 1200 residues, preferably between about 10
 and 100 residues and more preferably between about 25 and 35 residues.
 Such comparisons for substantial similarity are performed using
 methodology routine in the art.
 It is understood that minor modifications of primary amino acid sequence
 can result in an AGL8-like gene product that has substantially equivalent
 or enhanced function as compared to the AGL8 ortholog from which it was
 derived. Further, various molecules can be attached to an AGL8 ortholog or
 active segment thereof, for example, other polypeptides, antigenic or
 other peptide tags, carbohydrates, lipids, or chemical moieties. Such
 modifications are included within the term AGL8 ortholog as defined
 herein.
 One or more point mutations can be introduced into a nucleic acid molecule
 encoding an AGL8 ortholog to yield a modified nucleic acid molecule using,
 for example, site-directed mutagenesis (see Wu (Ed.), Meth. In Enzymol.
 Vol. 217, San Diego: Academic Press (1993); Higuchi, "Recombinant PCR" in
 Innis et al. (Ed.), PCR Protocols, San Diego: Academic Press, Inc. (1990),
 each of which is incorporated herein by reference). Such mutagenesis can
 be used to introduce a specific, desired amino acid insertion, deletion or
 substitution; alternatively, a nucleic acid sequence can be synthesized
 having random nucleotides at one or more predetermined positions to
 generate random amino acid substitutions. Scanning mutagenesis also can be
 useful in generating a modified nucleic acid molecule encoding
 substantially the amino acid sequence of an AGL8 ortholog.
 Modified nucleic acid molecules can be routinely assayed for the ability to
 alter normal development of the dehiscence zone and to delay seed
 dispersal. In the same manner as described in Examples I and III, a
 nucleic acid molecule encoding substantially the amino acid sequence of an
 AGL8 ortholog can be ectopically expressed, for example, using a
 constitutive regulatory element such as the CaMV 35S promoter or using a
 dehiscence zone-selective regulatory element such as the AGL1 promoter. If
 such ectopic expression results in a seed plant in which the dehiscence
 zone fails to develop and in which seed dispersal is delayed, the modified
 polypeptide or segment is an "AGL8 ortholog" as defined herein.
 A non-naturally occurring seed plant of the invention that is characterized
 by delayed seed dispersal can be one of a variety of seed plant species,
 such as a dehiscent seed plant or another monocotyledonous and
 dicotyledonous angiosperm or gymnosperm. A useful seed plant of the
 invention can be a dehiscent seed plant, and a particularly useful seed
 plant of the invention can be a member of the Brassicaceae, such as
 rapeseed, or a member of the Fabaceae, such as a soybean, pea, lentil or
 bean plant.
 As used herein, the term "seed plant" means an angiosperm or gymnosperm. An
 angiosperm is a seed-bearing plant whose seeds are borne in a mature ovary
 (fruit). An angiosperm commonly is recognized as a flowering plant.
 Angiosperms are divided into two broad classes based on the number of
 cotyledons, which are seed leaves that generally store or absorb food.
 Thus, a monocotyledonous angiosperm is an angiosperm having a single
 cotyledon, whereas a dicotyledonous angiosperm is an angiosperm having two
 cotyledons. A variety of angiosperms are known including, for example,
 oilseed plants, leguminous plants, fruit-bearing plants, ornamental
 flowers, cereal plants and hardwood trees, which general classes are not
 necessarily exclusive. The skilled artisan will recognize that the methods
 of the invention can be practiced using these or other angiosperms, as
 desired. A gymnosperm is a seed-bearing plant with seeds not enclosed in
 an ovary.
 In one embodiment, the invention provides a non-naturally occurring
 dehiscent seed plant that is characterized by delayed seed dispersal due
 to ectopic expression of a nucleic acid molecule encoding an AGL8-like
 gene product in the dehiscent seed plant. As used herein, the term
 "dehiscent seed plant" means a seed plant that produces a dry dehiscent
 fruit, which has fruit walls that open to permit escape of the seeds
 contained therein. Dehiscent fruits commonly contain several seeds and
 include the fruits known, for example, as legumes, capsules and siliques.
 In one embodiment, the invention provides a non-naturally occurring seed
 plant that is characterized by delayed seed dispersal due to ectopic
 expression of a nucleic acid molecule encoding an AGL8-like gene product,
 where the seed plant is a member of the Brassicaceae. The Brassicaceae,
 commonly known as the Brassicas, are a diverse group of crop plants with
 great economic value worldwide (see, for example, Williams and Hill,
 Science 232:1385-1389 (1986), which is incorporated herein by reference).
 The Brassicaceae produce seed oils for margarine, salad oil, cooking oil,
 plastic and industrial uses; condiment mustard; leafy, stored, processed
 and pickled vegetables; animal fodders and green manures for soil
 rejuvenation. A particularly useful non-naturally occurring Brassica seed
 plant of the invention is the oilseed plant canola.
 There are six major Brassica species of economic importance, each
 containing a range of plant forms. Brassica napus includes plants such as
 the oilseed rapes and rutabaga. Brassica oleracea are the cole crops such
 as cabbage, cauliflower, kale, kohlrabi and Brussels sprouts. Brassica
 campestris (Brassica rapa) includes plants such as Chinese cabbage, turnip
 and pak choi. Brassica juncea includes a variety of mustards; Brassica
 nigra is the black mustard; and Brassica carinata is Ethiopian mustard.
 The skilled artisan understands that any member of the Brassicaceae can be
 modified as disclosed herein to produce a non-naturally occurring Brassica
 plant characterized by delayed seed dispersal.
 In a second embodiment, the invention provides a non-naturally occurring
 seed plant that is characterized by delayed seed dispersal due to ectopic
 expression of a nucleic acid molecule encoding an AGL8-like gene product,
 where the seed plant is a member of the Fabaceae. The Fabaceae, which are
 commonly known as members of the pea family, are seed plants that produce
 a characteristic dry dehiscent fruit known as a legume. The legume is
 derived from a single carpel and dehisces along the suture of the carpel
 margins and along the median vein. The Fabaceae encompass both grain
 legumes and forage legumes. Grain legumes include, for example, soybean
 (glycine), pea, chickpea, moth bean, broad bean, kidney bean, lima bean,
 lentil, cowpea, dry bean and peanut. Forage legumes include alfalfa,
 lucerne, birdsfoot trefoil, clover, stylosanthes species, lotononis
 bainessii and sainfoin. The skilled artisan will recognize that any member
 of the Fabaceae can be modified as disclosed herein to produce a
 non-naturally occurring seed plant of the invention characterized by
 delayed seed dispersal.
 A non-naturally occurring seed plant of the invention characterized by
 delayed seed dispersal also can be a member of the plant genus Cuphea
 (family Lythraceae). A Cuphea seed plant is particularly valuable since
 Cuphea oilseeds contain industrially and nutritionally important
 medium-chain fatty acids, especially lauric acid, which is currently
 supplied only by coconut and palm kernel oils.
 A non-naturally occurring seed plant of the invention also can be, for
 example, one of the monocotyledonous grasses, which produce many of the
 valuable small-grain cereal crops of the world. In a non-naturally
 occurring small grain cereal plant of the invention, grain remains on the
 seed plant longer and, Ectopic expression of a nucleic acid molecule
 encoding an AGL8-like gene product, or suppression of AGL1 and AGL5
 expression as described below, can be useful in generating a non-naturally
 occurring small grain cereal plant, such as a barley, wheat, oat, rye,
 orchard grass, guinea grass, sorghum or turf grass plant characterized by
 delayed seed dispersal.
 The invention also provides a transgenic seed plant that is characterized
 by delayed seed dispersal due to ectopic expression of a nucleic acid
 molecule encoding an AGL8-like gene product. In a transgenic seed plant of
 the invention, the ectopically expressed nucleic acid molecule encoding an
 AGL8-like gene product can be operatively linked to an exogenous
 regulatory element. The invention provides, for example, a transgenic seed
 plant characterized by delayed seed dispersal having an ectopically
 expressed nucleic acid molecule encoding an AGL8-like gene product that is
 operatively linked to an exogenous constitutive regulatory element. In one
 embodiment, the invention provides a transgenic seed plant that is
 characterized by delayed seed dispersal due to ectopic expression of an
 exogenous nucleic acid molecule encoding substantially the amino acid
 sequence of an AGL8 ortholog operatively linked to an exogenous
 cauliflower mosaic virus 35S promoter.
 The invention also provides a transgenic seed plant that is characterized
 by delayed seed dispersal due to ectopic expression of a nucleic acid
 molecule encoding an AGL8-like gene product operatively linked to a
 dehiscence zone-selective regulatory element. The dehiscence
 zone-selective regulatory element can be, for example, an AGL1 regulatory
 element or AGL5 regulatory element. The AGL1 regulatory element can be
 derived from the Arabidopsis AGL1 genomic sequence disclosed herein as SEQ
 ID NO:3 and can be, for example, a 5' regulatory sequence or intronic
 regulatory element. Similarly, the AGL5 regulatory element can be derived
 from the Arabidopsis AGL5 genomic sequence disclosed herein as SEQ ID NO:4
 and can be, for example, a 5' regulatory sequence or intronic regulatory
 element
 In one embodiment, a transgenic seed plant of the invention has an
 ectopically expressed exogenous nucleic acid molecule encoding
 substantially the amino acid sequence of an AGL8 ortholog operatively
 linked to a dehiscence zone-selective regulatory element that is an AGL1
 regulatory element having at least fifteen contiguous nucleotides of
 nucleotides 1 to 2599 of SEQ ID NO:3; nucleotides 2833 to 4128 of SEQ ID
 NO:3; nucleotides 4211 to 4363 of SEQ ID NO:3; nucleotides 4426 to 4554 of
 SEQ ID NO:3; nucleotides 4796 to 4878 of SEQ ID NO:3; nucleotides 4921 to
 5028 of SEQ ID NO:3; or nucleotides 5421 to 5682 of SEQ ID NO:3.
 In another embodiment, a transgenic seed plant of the invention has an
 ectopically expressed exogenous nucleic acid molecule encoding
 substantially the amino acid sequence of an AGL8 ortholog operatively
 linked to a dehiscence zone-selective regulatory element that is an AGL5
 regulatory element having at least fifteen contiguous nucleotides of
 nucleotides 1 to 1890 of SEQ ID NO:4; nucleotides 2536 to 2683 of SEQ ID
 NO:4; nucleotides 2928 to 5002 of SEQ ID NO:4; nucleotides 5085 to 5204 of
 SEQ ID NO:4; nucleotides 5367 to 5453 of SEQ ID NO:4; nucleotides 5645 to
 5734 of SEQ ID NO:4; or nucleotides 6062 to 6138 of SEQ ID NO:4.
 As used herein, the term "transgenic" refers to a seed plant that contains
 an exogenous nucleic acid molecule, which can be derived from the same
 seed plant species or a heterologous seed plant species.
 The term "exogenous," as used herein in reference to a nucleic acid
 molecule and a transgenic seed plant, means a nucleic acid molecule
 originating from outside the seed plant. An exogenous nucleic acid
 molecule can be, for example, a nucleic acid molecule encoding an
 AGL8-like gene product or an exogenous regulatory element such as a
 constitutive regulatory element or a dehiscence zone-selective regulatory
 element, as described further below. An exogenous nucleic acid molecule
 can have a naturally occurring or non-naturally occurring nucleotide
 sequence and can be a heterologous nucleic acid molecule derived from a
 different seed plant species than the seed plant into which the nucleic
 acid molecule is introduced or can be a nucleic acid molecule derived from
 the same seed plant species as the seed plant into which it is introduced.
 The term "operatively linked," as used in reference to a regulatory element
 and a nucleic acid molecule, means that the regulatory element confers
 regulated expression upon the operatively linked nucleic acid molecule.
 Thus, the term "operatively linked," as used in reference to an exogenous
 regulatory element such as a dehiscence zone-selective regulatory element
 and a nucleic acid molecule encoding an AGL8-like gene product, means that
 the dehiscence zone-selective regulatory element is linked to the nucleic
 acid molecule encoding an AGL8-like gene product such that the expression
 pattern of the dehiscence zone-selective regulatory element is conferred
 upon the nucleic acid molecule encoding the AGL8-like gene product. It is
 recognized that a regulatory element and a nucleic acid molecule that are
 operatively linked have, at a minimum, all elements essential for
 transcription, including, for example, a TATA box.
 As used herein, the term "constitutive regulatory element" means a
 regulatory element that confers a level of expression upon an operatively
 linked nucleic molecule that is relatively independent of the cell or
 tissue type in which the constitutive regulatory element is expressed. A
 constitutive regulatory element that is expressed in a seed plant
 generally is widely expressed in a large number of cell and tissue types.
 A variety of constitutive regulatory elements useful for ectopic expression
 in a transgenic seed plant are well known in the art. The cauliflower
 mosaic virus 35S (CaMV 35S) promoter, for example, is a well-characterized
 constitutive regulatory element that produces a high level of expression
 in all plant tissues (Odell et al., Nature 313:810-812 (1985)). The CaMV
 35S promoter can be particularly useful due to its activity in numerous
 diverse seed plant species (Benfey and Chua, Science 250:959-966 (1990);
 Futterer et al., Physiol. Plant 79:154 (1990); Odell et al., supra, 1985).
 A tandem 35S promoter, in which the intrinsic promoter element has been
 duplicated, confers higher expression levels in comparison to the
 unmodified 35S promoter (Kay et al., Science 236:1299 (1987)). Other
 constitutive regulatory elements useful for ectopically expressing a
 nucleic acid molecule encoding an AGL8-like gene product in a transgenic
 seed plant of the invention include, for example, the cauliflower mosaic
 virus 19S promoter; the Figwort mosaic virus promoter; and the nopaline
 synthase (nos) gene promoter (Singer et al., Plant Mol. Biol. 14:433
 (1990); An, Plant Physiol. 81:86 (1986)).
 Additional constitutive regulatory elements including those for efficient
 ectopic expression in monocots also are known in the art, for example, the
 pEmu promoter and promoters based on the rice Actin-1 5' region (Last et
 al., Theor. Appl. Genet. 81:581 (1991); Mcelroy et al., Mol. Gen. Genet.
 231:150 (1991); Mcelroy et al., Plant Cell 2:163 (1990)). Chimeric
 regulatory elements, which combine elements from different genes, also can
 be useful for ectopically expressing a nucleic acid molecule encoding an
 AGL8-like gene product (Comai et al., Plant Mol. Biol. 15:373 (1990)). One
 skilled in the art understands that a particular constitutive regulatory
 element is chosen based, in part, on the seed plant species in which a
 nucleic acid molecule encoding an AGL8-like gene product is to be
 ectopically expressed and on the desired level of expression.
 An exogenous regulatory element useful in a transgenic seed plant of the
 invention also can be an inducible regulatory element, which is a
 regulatory element that confers conditional expression upon an operatively
 linked nucleic acid molecule, where expression of the operatively linked
 nucleic acid molecule is increased in the presence of a particular
 inducing agent or stimulus as compared to expression of the nucleic acid
 molecule in the absence of the inducing agent or stimulus. Particularly
 useful inducible regulatory elements include copper-inducible regulatory
 elements (Mett et al., Proc. Natl. Acad. Sci. USA 90:4567-4571 (1993);
 Furst et al., Cell 55:705-717 (1988)); tetracycline and
 chlor-tetracycline-inducible regulatory elements (Gatz et al., Plant J.
 2:397-404 (1992); Roder et al., Mol. Gen. Genet. 243:32-38 (1994); Gatz,
 Meth. Cell Biol. 50:411-424 (1995)); ecdysone inducible regulatory
 elements (Christopherson et al., Proc. Natl. Acad. Sci. USA 89:6314-6318
 (1992); Kreutzweiser et al., Ecotoxicol. Environ. Safety 28:14-24 (1994));
 heat shock inducible regulatory elements (Takahashi et al., Plant Physiol.
 99:383-390 (1992); Yabe et al., Plant Cell Physiol. 35:1207-1219 (1994);
 Ueda et al., Mol. Gen. Genet. 250:533-539 (1996)); and lac operon
 elements, which are used in combination with a constitutively expressed
 lac repressor to confer, for example, IPTG-inducible expression (Wilde et
 al., EMBO J. 11:1251-1259 (1992)).
 An inducible regulatory element useful in the transgenic seed plants of the
 invention also can be, for example, a nitrate-inducible promoter derived
 from the spinach nitrite reductase gene (Back et al., Plant Mol. Biol.
 17:9 (1991)) or a light-inducible promoter, such as that associated with
 the small subunit of RuBP carboxylase or the LHCP gene families (Feinbaum
 et al., Mol. Gen. Genet. 226:449 (1991); Lam and Chua, Science 248:471
 (1990)). Additional inducible regulatory elements include salicylic acid
 inducible regulatory elements (Uknes et al., Plant Cell 5:159-169 (1993);
 Bi et al., Plant J. 8:235-245 (1995)); plant hormone-inducible regulatory
 elements (Yamaguchi-Shinozaki et al., Plant Mol. Biol. 15:905 (1990);
 Kares et al., Plant Mol. Biol. 15:225 (1990)); and human hormone-inducible
 regulatory elements such as the human glucocorticoid response element
 (Schena et al., Proc. Natl. Acad. Sci. USA 88:10421 (1991)).
 It should be recognized that a non-naturally occurring seed plant of the
 invention, which contains an ectopically expressed nucleic acid molecule
 encoding an AGL8-like gene product, also can contain one or more
 additional modifications, including naturally and non-naturally occurring
 modifications, that can modulate the delay in seed dispersal. For example,
 the plant hormone ethylene promotes fruit dehiscence, and modified
 expression or activity of positive or negative regulators of the ethylene
 response can be included in a seed plant of the invention (see, generally,
 Meakin and Roberts, J. Exp. Botany 41:1003-1011 (1990); Ecker, Science
 268:667-675 (1995); Chao et al., Cell 89:1133-1144 (1997)).
 Mutations in positive regulators of the ethylene response show a reduction
 or absence of responsiveness to treatment with exogenous ethylene.
 Arabidopsis mutations in positive regulators of the ethylene response
 include mutations in etr, which inactivate a histidine kinase ethylene
 receptor (Bleeker et al., Science 241:1086-1089 (1988); Schaller and
 Bleeker, Science 270:1809-1811 (1995)); ers (Hua et al., Science
 269:1712-1714 (1995)); ein2 (Guzman and Ecker, Plant Cell 2:513 (1990));
 ein3 (Rothenberg and Ecker, Sem. Dev. Biol. Plant Dev. Genet. 4:3-13
 (1993); Kieber and Ecker, Trends Genet. 9:356-362 (1993)); ain1 (van der
 Straeten et al., Plant Physiol. 102:401-408 (1993)); eti (Harpham et al.,
 An. Bot. 68:55 (1991)) and ein4, ein5, ein6, and ein7 (Roman et al.,
 Genetics 139: 1393-1409 (1995)). Similar genetic functions are found in
 other seed plant species; for example, the never-ripe mutation corresponds
 to etr and confers ethylene insensitivity in tomato (Lanahan et al., The
 Plant Cell 6:521-530 (1994); Wilkinson et al., Science 270:1807-1809
 (1995)). A seed plant of the invention can include a modification that
 results in altered expression or activity of any such positive regulator
 of the ethylene response. A mutation in a positive regulator, for example,
 can be included in a seed plant of the invention and can modify the delay
 in seed dispersal in such plants, for example, by further postponing the
 delay in seed dispersal.
 Mutations in negative regulators of the ethylene response display ethylene
 responsiveness in the absence of exogenous ethylene. Such mutations
 include those relating to ethylene overproduction, for example, the eto1,
 eto2, and eto3 mutants, and those relating to constitutive activation of
 the ethylene signalling pathway, for example, mutations in CTR1, a
 negative regulator with sequence similarity to the Raf family of protein
 kinases (Kieber et al., Cell 72:427-441 (1993), which is incorporated
 herein by reference). A seed plant of the invention can include a
 modification that results in altered expression or activity of any such
 negative regulator of the ethylene response. A mutation resulting in
 ethylene responsiveness in the absence of exogenous ethylene, for example,
 can be included in a non-naturally occurring seed plant of the invention
 and can modify, for example, diminish, the delay in seed dispersal.
 Fruit morphological mutations also can be included in a seed plant of the
 invention. Such mutations include those in carpel identity genes such as
 AGAMOUS (Bowman et al., supra, 1989; Yanofsky et al., supra, 1990) and in
 genes required for normal fruit development such as ETTIN, CRABS CLAW,
 SPATULA, AGL8 and TOUSLED (Sessions et al., Development 121:1519-1532
 (1995); Alvarez and Smyth, Flowering Newsletter 23:12-17 (1997); and Roe
 et al., Cell 75:939-950 (1993)). Thus, it is understood that a seed plant
 of the invention having an ectopically expressed nucleic acid molecule
 encoding an AGL8-like gene product can include one or more additional
 genetic modifications, which can diminish or enhance the delay in seed
 dispersal.
 The present invention also provides methods of producing a non-naturally
 occurring seed plant characterized by delayed seed dispersal. A method of
 the invention entails ectopically expressing a nucleic acid molecule
 encoding an AGL8-like gene product in the seed plant, whereby seed
 dispersal is delayed due to ectopic expression of the nucleic acid
 molecule.
 As discussed above, the term "ectopically" refers to expression of a
 nucleic acid molecule encoding an AGL8-like gene product in a cell type
 other than a cell type in which the nucleic acid molecule is normally
 expressed, at a time other than a time at which the nucleic acid molecule
 is normally expressed or at n expression level other than the level at
 which the nucleic acid normally is expressed. In wild type Arabidopsis,
 for example, AGL8 expression is normally restricted during the later
 stages of floral development to the carpel valves and is not seen in the
 outer replum. In the methods of the invention, particularly useful ectopic
 expression of a nucleic acid molecule encoding an AGL8-like gene product
 involves expression in the cells of the outer replum, which are the
 progenitors of the dehiscence zone.
 Actual ectopic expression of an AGL8-like gene product is dependent on
 various factors. The ectopic expression can be widespread expression
 throughout most or all plant tissues or can be expression restricted to a
 small number of plant tissues, and can be achieved by a variety of routine
 techniques. Mutagenesis, including seed or pollen mutagenesis, can be used
 to generate a non-naturally occurring seed plant, in which a nucleic acid
 molecule encoding an AGL8-like gene product is ectopically expressed.
 Ethylmethane sulfonate (EMS) mutagenesis, transposon mediated mutagenesis
 or T-DNA mediated mutagenesis also can be useful in ectopically expressing
 an AGL8-like gene product to produce a seed plant characterized by delayed
 seed dispersal (see, generally, Glick and Thompson, supra, 1993). While
 not wishing to be bound by any particular mechanism, ectopic expression in
 a mutagenized plant can result from inactivation of one or more negative
 regulators of AGL8, for example, from the combined inactivation of AGL1
 and AGL5.
 Ectopic expression of an AGL8-like gene product also can be achieved by
 expression of a nucleic acid encoding an AGL8-like gene product from a
 heterologous regulatory element or from a modified variant of its own
 promoter. Heterologous regulatory elements include constitutive regulatory
 elements, which result in expression of the AGL8-like gene product in the
 outer replum as well as in a variety of other cell types, and dehiscence
 zone-selective regulatory elements, which produce selective expression of
 an AGL8-like gene product in a limited number of cell types including the
 cells of the valve margin or the dehiscence zone.
 Ectopic expression of a nucleic acid molecule encoding an AGL8-like gene
 product can be achieved using an endogenous or exogenous nucleic acid
 molecule encoding an AGL8-like gene product. A recombinant exogenous
 nucleic acid molecule can contain a heterologous regulatory element that
 is operatively linked to a nucleic acid sequence encoding an AGL8-like
 gene product. Methods for producing the desired recombinant nucleic acid
 molecule under control of a heterologous regulatory element and for
 producing a non-naturally occurring seed plant of the invention are well
 known in the art (see, generally, Sambrook et al., supra, 1989; Glick and
 Thompson, supra, 1993).
 An exogenous nucleic acid molecule can be introduced into a seed plant for
 ectopic expression using a variety of transformation methodologies
 including Agrobacterium-mediated transformation and direct gene transfer
 methods such as electroporation and microprojectile-mediated
 transformation (see, generally, Wang et al. (eds), Transformation of
 Plants and Soil Microorganisms, Cambridge, UK: University Press (1995),
 which is incorporated herein by reference). Transformation methods based
 upon the soil bacterium Agrobacterium tumefaciens are particularly useful
 for introducing an exogenous nucleic acid molecule into a seed plant. The
 wild type form of Agrobacterium contains a Ti (tumor-inducing) plasmid
 that directs production of tumorigenic crown gall growth on host plants.
 Transfer of the tumor-inducing T-DNA region of the Ti plasmid to a plant
 genome requires the Ti plasmid-encoded virulence genes as well as T-DNA
 borders, which are a set of direct DNA repeats that delineate the region
 to be transferred. An Agrobacterium-based vector is a modified form of a
 Ti plasmid, in which the tumor inducing functions are replaced by the
 nucleic acid sequence of interest to be introduced into the plant host.
 Agrobacterium-mediated transformation generally employs cointegrate vectors
 or, preferably, binary vector systems, in which the components of the Ti
 plasmid are divided between a helper vector, which resides permanently in
 the Agrobacterium host and carries the virulence genes, and a shuttle
 vector, which contains the gene of interest bounded by T-DNA sequences. A
 variety of binary vectors are well known in the art and are commercially
 available, for example, from Clontech (Palo Alto, Calif.). Methods of
 coculturing Agrobacterium with cultured plant cells or wounded tissue such
 as leaf tissue, root explants, hypocotyledons, stem pieces or tubers, for
 example, also are well known in the art (Glick and Thompson, supra, 1993).
 Wounded cells within the plant tissue that have been infected by
 Agrobacterium can develop organs de novo when cultured under the
 appropriate conditions; the resulting transgenic shoots eventually give
 rise to transgenic plants that ectopically express a nucleic acid molecule
 encoding an AGL8-like gene product. Agrobacteriurn also can be used for
 transformation of whole seed plants as described in Bechtold et al., C.R.
 Acad. Sci. Paris, Life Sci. 316:1194-1199 (1993), which is incorporated
 herein by reference). Agrobacterium-mediated transformation is useful for
 producing a variety of transgenic seed plants (Wang et al., supra, 1995)
 including transgenic plants of the Brassicaceae family, such as rapeseed,
 Arabidopsis, mustard, and flax, and transgenic plants of the Fabaceae
 family such as soybean, pea, lentil and bean.
 Microprojectile-mediated transformation also can be used to produce a
 transgenic seed plant that ectopically expresses an AGL8-like gene
 product. This method, first described by Klein et al. (Nature 327:70-73
 (1987), which is incorporated herein by reference), relies on
 microprojectiles such as gold or tungsten that are coated with the desired
 nucleic acid molecule by precipitation with calcium chloride, spermidine
 or PEG. The microprojectile particles are accelerated at high speed into
 an angiosperm tissue using a device such as the BIOLISTIC PD-1000 (Biorad;
 Hercules Calif.).
 Microprojectile-mediated delivery or "particle bombardment" is especially
 useful to transform seed plants that are difficult to transform or
 regenerate using other methods. Microprojectile-mediated transformation
 has been used, for example, to generate a variety of transgenic plant
 species, including cotton, tobacco, corn, hybrid poplar and papaya (see
 Glick and Thompson, supra, 1993) as well as cereal crops such as wheat,
 oat, barley, sorghum and rice (Duan et al., Nature Biotech. 14:494-498
 (1996); Shimamoto, Curr. Opin. Biotech. 5:158-162 (1994), each of which is
 incorporated herein by reference). In view of the above, the skilled
 artisan will recognize that Agrobacterium-mediated or
 microprojectile-mediated transformation, as disclosed herein, or other
 methods known in the art can be used to introduce a nucleic acid molecule
 encoding an AGL8-like gene product into a seed plant for ectopic
 expression.
 In another embodiment, the invention provides a non-naturally occurring
 seed plant that is characterized by delayed seed dispersal due to
 suppression of both AGL1 expression and AGL5 expression in the seed plant.
 Such a non-naturally occurring seed plant characterized by delayed seed
 dispersal can be, for example, an agl1 agl5 double mutant.
 As disclosed herein, loss-of-function mutations in the AGL1 and AGL5 genes
 were produced by a combination of homologous recombination and disruptive
 T-DNA insertion (see Example II). Neither AGL1 nor AGL5 RNA was expressed
 in the resulting agl1 agl5 double mutant, and scanning electron microscopy
 revealed that the dehiscence zone failed to develop normally in these
 mutant seed plants. Furthermore, the mature fruits of these seed plants
 failed to undergo dehiscence, as shown in FIG. 5. These results indicate
 that AGL1 or AGL5 gene expression is required for normal development of
 the dehiscence zone and that suppression of AGL1 expression combined with
 suppression of AGL5 expression in the seed plant can delay dehiscence,
 allowing the process of pod shatter to be controlled.
 The Arabidopsis AGL1 and AGL5 genes encode MADS box proteins with 85%
 identity at the amino acid level (see Tables 1 and 2). The AGL1 and AGL5
 RNA expression patterns also are strikingly similar. In particular, both
 RNAs are specifically expressed in flowers, where they accumulate in
 developing carpels. In particular, strong expression of these genes is
 observed in the outer replum along the valve/replum boundary (Ma et al.,
 supra, 1991; Savidge et al., The Plant Cell 7:721-723 (1995); Flanagan et
 al., The Plant Journal 10:343-353 (1996), each of which is incorporated
 herein by reference). Thus, AGL1 and AGL5 are expressed in the valve
 margin, at least within the cells of the outer replum.
 TABLE 1
 Amino acid identity in the MADS domain and
 K-domain of AGAMOUS, AGL1 and AGL5
 AGAMOUS AGL1 AGL5
 MADS K MADS K MADS K
 AGAMOUS -- -- 95% 68% 95% 62%
 AGL1 -- -- -- -- 100% 92%
 AGL5 -- -- -- -- -- --
 TABLE 1
 Amino acid identity in the MADS domain and
 K-domain of AGAMOUS, AGL1 and AGL5
 AGAMOUS AGL1 AGL5
 MADS K MADS K MADS K
 AGAMOUS -- -- 95% 68% 95% 62%
 AGL1 -- -- -- -- 100% 92%
 AGL5 -- -- -- -- -- --
 As used herein, the term "AGL1" refers to Arabidopsis AGL1 (SEQ ID NO:6) or
 an ortholog of Arabidopsis AGL1 (SEQ ID NO:6). An AGL1 ortholog is a MADS
 box gene product expressed, at least in part, in the valve margins of a
 seed plant and having homology to the amino acid sequence of Arabidopsis
 AGL1 (SEQ ID NO:6). AGL1 or an AGL1 ortholog can function, in part, by
 forming a complex with an AGL8-like gene product. An AGL1 ortholog
 generally has an amino acid sequence having at least about 63% amino acid
 identity with Arabidopsis AGL1 (SEQ ID NO:6) and includes polypeptides
 having greater than about 70%, 75%, 85% or 95% amino acid identity with
 Arabidopsis AGL1 (SEQ ID NO:6). Given the close relatedness of the AGL1
 and AGL5 gene products, one skilled in the art will recognize that an AGL1
 ortholog can be distinguished from an AGL5 ortholog by being more closely
 related to Arabidopsis AGL1 (SEQ ID NO:6) than to Arabidopsis AGL5 (SEQ ID
 NO:8). An AGL1 ortholog can function in wild type plants, like Arabidopsis
 AGL1, to limit the domain of AGL8-like gene product expression to the
 carpel valves during the later stages of floral development.
 As used herein, the term "AGL5" refers to Arabidopsis AGL5 (SEQ ID NO:8) or
 to an ortholog of Arabidopsis AGL5 (SEQ ID NO:8). An AGL5 ortholog is a
 MADS box gene product expressed, at least in part, in the valve margins of
 a seed plant and having homology to the amino acid sequence of Arabidopsis
 AGL5 (SEQ ID NO:8). AGL5 or an AGL5 ortholog can function, in part, by
 forming a complex with an AGL8-like gene product as shown in Example IV.
 An AGL5 ortholog generally has an amino acid sequence having at least
 about 60% amino acid identity with Arabidopsis AGL5 (SEQ ID NO:8) and
 includes polypeptides having greater than about 65%, 70%, 75%, 85% or 95%
 amino acid identity with Arabidopsis AGL5 (SEQ ID NO:8). Given the close
 relatedness of the AGL1 and AGL5 gene products, one skilled in the art
 will recognize that an AGL5 ortholog can be distinguished from an AGL1
 ortholog by being more closely related to Arabidopsis AGL5 (SEQ ID NO:8)
 than to Arabidopsis AGL1 (SEQ ID NO:6). An AGL5 ortholog can function in
 wild type plants, like Arabidopsis AGL5, to limit the domain of AGL8-like
 gene product expression to the carpel valves during the later stages of
 floral development.
 The term "suppressed," as used herein in reference to AGL1 expression,
 means that the amount of functional AGL1 protein is reduced in a seed
 plant in comparison with the amount of functional AGL1 protein in the
 corresponding wild type seed plant. Similarly, when used in reference to
 AGL5 expression, the term suppressed means that the amount of functional
 AGL5 protein is reduced in a seed plant in comparison with the amount of
 functional AGL5 protein in the corresponding wild type seed plant. Thus,
 the term "suppressed," as used herein, encompasses the absence of AGL1 or
 AGL5 protein in a seed plant, as well as protein expression that is
 present but reduced as compared to the level of AGL1 or AGL5 protein
 expression in a wild type seed plant. Furthermore, the term suppressed
 refers to AGL1 or AGL5 protein expression that is reduced throughout the
 entire domain of AGL1 or AGL5 expression, or to expression that is reduced
 in some part of the AGL1 or AGL5 expression domain, provided that the
 resulting seed plant is characterized by delayed seed dispersal.
 As used herein, the term "suppressed" also encompasses an amount of AGL1 or
 AGL5 protein that is equivalent to wild type AGL1 or AGL5 expression, but
 where the AGL1 or AGL5 protein has a reduced level of activity. As
 discussed above, AGL1 and AGL5 each contain a conserved MADS domain; point
 mutations or gross deletions within the MADS domain that reduce the
 DNA-binding activity of AGL1 or AGL5 can reduce or destroy the activity of
 AGL1 or AGL5 and, therefore, "suppress" AGL1 or AGL5 expression as defined
 herein. One skilled in the art will recognize that, preferably, AGL1
 expression is essentially absent in the valve margin of a seed plant or
 the AGL1 protein is essentially non-functional and, similarly, that,
 preferably, AGL5 expression is essentially absent in the valve margin of
 the seed plant or the AGL5 protein is essentially non-functional.
 A variety of methodologies can be used to suppress AGL1 or AGL5 expression
 in a seed plant. Suppression can be achieved by directly modifying the
 AGL1 or AGL5 genomic locus, for example, by modifying an AGL1 or AGL5
 regulatory sequence such that transcription or translation from the AGL1
 or AGL5 locus is reduced, or by modifying an AGL1 or AGL5 coding sequence
 such that non-functional AGL1 or AGL5 protein is produced. Suppression of
 AGL1 or AGL5 expression in a seed plant also can be achieved indirectly,
 for example, by modifying the expression or activity of a protein that
 regulates AGL1 or AGL5 expression. Methodologies for effecting suppression
 of AGL1 or AGL5 expression in a seed plant include, for example,
 homologous recombination, chemical and transposon-mediated mutagenesis,
 cosuppression and antisense-based techniques and dominant negative
 methodologies.
 Homologous recombination of AGL1 or AGL5 can be used to suppress AGL1 or
 AGL5 expression in a seed plant as described in Kempin et al., Nature
 389:802-803 (1997), which is incorporated herein by reference. Homologous
 recombination can be used, for example, to replace the wild type AGL5
 genomic sequence with a construct in which the gene for kanamycin
 resistance is flanked by at least about 1 kb of AGL5 sequence. The use of
 homologous recombination to suppress AGL5 expression is set forth in
 Example II.
 Suppression of AGL1 or AGL5 expression also can be achieved by producing a
 loss-of-function mutation using transposon-mediated insertional
 mutagenesis with Ds transposons or Stm transposons (see, for example,
 Sundaresan et al., Genes Devel. 9:1797-1810 (1995), which is incorporated
 herein by reference). Insertion of a transposon into an AGL1 or AGL5
 target gene can be identified, for example, by restriction mapping, which
 can identify the presence of an insertion in the gene promoter or in the
 coding region, such that expression of functional gene product is
 suppressed. Insertion of a transposon also can be identified by detecting
 an absence of the mRNA encoded by the target gene or by the detecting the
 absence of the gene product in valve margin. Suppression of AGL1 or AGL5
 expression also can be achieved by producing a loss-of-function mutation
 using T-DNA-mediated insertional mutagenesis (see Krysan et al., Proc.
 Natl. Acad. Sci., USA 93:8145-8150 (1996)). The use of T-DNA-mediated
 insertional mutagenesis to suppress AGL1 expression is disclosed in
 Example II.
 Suppression of AGL1 or AGL5 expression in a seed plant also can be achieved
 using cosuppression, which is a well known methodology that relies on
 expression of a nucleic acid molecule in the sense orientation to produce
 coordinate silencing of the introduced nucleic acid molecule and the
 homologous endogenous gene (see, for example, Flavell, Proc. Natl. Acad.
 Sci., USA 91:3490-3496 (1994); Kooter and Mol, Current Opin. Biol.
 4:166-171 (1993), each of which is incorporated herein by reference).
 Cosuppression is induced most strongly by a large number of transgene
 copies or by overexpression of transgene RNA and can be enhanced by
 modification of the transgene such that it fails to be translated.
 Antisense nucleic acid molecules encoding AGL1 and AGL5 gene products, or
 fragments thereof, also can be used to suppress expression of AGL1 and
 AGL5 in a seed plant. Antisense nucleic acid molecules reduce mRNA
 translation or increase mRNA degradation, thereby suppressing gene
 expression (see, for example, Kooter and Mol, supra, 1993; Pnueli et al.,
 The Plant Cell Vol. 6, 175-186 (1994), which is incorporated herein by
 reference).
 To produce a non-naturally occurring seed plant of the invention, in which
 AGL1 and AGL5 expression each are suppressed, the one or more sense or
 antisense nucleic acid molecules can be expressed under control of a
 strong regulatory element that is expressed, at least in part, in the
 valve margin of the seed plant. The constitutive CaMV 35S promoter (Odell
 et al., supra, 1985), for example, or other constitutive promoters as
 disclosed herein, can be useful in the methods of the invention.
 Dehiscence zone-selective regulatory elements also can be useful for
 expressing one or more sense or antisense nucleic acid molecules in order
 to suppress AGL1 and AGL5 expression in a seed plant
 The skilled artisan will recognize that effective suppression of endogenous
 AGL1 and AGL5 gene expression depends upon the one or more introduced
 nucleic acid molecules having a high percentage of homology with the
 corresponding endogenous gene loci. Nucleic acid molecules encoding
 Arabidopsis AGL1 (SEQ ID NO:5) and AGL5 (SEQ ID NO:7) are provided herein
 (see, also, Ma et al., supra, 1991). Nucleic acid molecules encoding
 Arabidopsis AGL1 and AGL5 can be useful in the methods of the invention or
 for isolating orthologous AGL1 and AGL5 sequences.
 The homology requirement for effective suppression using homologous
 recombination, cosuppression or antisense methodology can be determined
 empirically. In general, a minimum of about 80-90% nucleic acid sequence
 identity is preferred for effective suppression of AGL1 or AGL5
 expression. Thus, a nucleic acid molecule encoding a gene ortholog from
 the family or genus of the seed plant species into which the nucleic acid
 molecule is to be introduced is preferred for generating the non-naturally
 occurring seed plants of the invention using homologous recombination,
 cosuppression or antisense technology. More preferably, a nucleic acid
 molecule encoding a gene ortholog from the same seed plant species is used
 for suppressing AGL1 expression and AGL5 expression in a seed plant of the
 invention. For example, nucleic acid molecules encoding canola AGL1 and
 AGL5 are preferable for suppressing AGL1 and AGL5 expression in a canola
 plant.
 Although use of a highly homologous nucleic acid molecule is preferred in
 the methods of the invention, the nucleic acid molecule to be used for
 homologous recombination, cosuppression or antisense suppression need not
 contain in its entirety the AGL1 or AGL5 sequence to be suppressed. Thus,
 a sense or antisense nucleic acid molecule encoding only a portion of
 Arabidopsis AGL1 (SEQ ID NO:5), for example, or a sense or antisense
 nucleic acid molecule encoding only a portion of Arabidopsis AGL5 (SEQ ID
 NO:7) can be useful for producing a non-naturally occurring seed plant of
 the invention, in which AGL1 and AGL5 expression each are suppressed.
 A portion of a nucleic acid molecule to be homologously recombined with an
 AGL1 or AGL5 locus generally contains at least about 1 kb of sequence
 homologous to the targeted gene and preferably contains at least about 2
 kb, more preferably at least about 3 kb and can contain at least about 5
 kb of sequence homologous to the targeted gene. A portion of a nucleic
 acid molecule encoding an AGL1 or AGL5 to be used for cosuppression or
 antisense suppression generally contains at least about 50 base pairs to
 the full-length of the nucleic acid molecule encoding the AGL1 or AGL5
 ortholog. In contrast to an active segment, as defined herein, a portion
 of a nucleic acid molecule to be used for homologous recombination,
 cosuppression or antisense suppression need not encode a functional part
 of a gene product.
 A dominant negative construct also can be used to suppress AGL1 or AGL5
 expression in a seed plant. A dominant negative construct useful in the
 invention generally contains a portion of the complete AGL1 or AGL5 coding
 sequence sufficient, for example, for DNA-binding or for a protein-protein
 interaction such as a homodimeric or heterodimeric protein-protein
 interaction but lacking the transcriptional activity of the wild type
 protein. For example, a carboxy-terminal deletion mutant of AGAMOUS was
 used as a dominant negative construct to suppress expression of the MADS
 box gene AGAMOUS (Mizukami et al., Plant Cell 8:831-844 (1996), which is
 incorporated by reference herein). One skilled in the art understands
 that, similarly, a dominant negative AGL1 or AGL5 construct can be used to
 suppress AGL1 or AGL5 expression in a seed plant. A useful dominant
 negative construct can be a deletion mutant encoding, for example, the
 MADS box domain alone ("M"), the MADS box domain and "intervening" region
 ("MI"); the MADS box, "intervening" and "K" domains ("MIK"); or the
 "intervening," "K" and carboxy-terminal domains ("IKC").
 In a preferred embodiment, a non-naturally occurring seed plant of the
 invention is an agl1 agl5 double mutant. An agl1 agl5 double mutant is a
 particularly useful non-naturally occurring seed plant that is
 characterized by delayed seed dispersal.
 As used herein, the term "agl1 agl5 double mutant" means a seed plant
 having a loss-of-function mutation at the AGL1 locus and a
 loss-of-function mutation at the AGL5 locus. Loss-of-function mutations
 encompass point mutations, including substitutions, deletions and
 insertions, as well as gross modifications of an AGL1 and AGL5 locus and
 can be located in coding or non-coding sequences. One skilled in the art
 understands that any such loss-of-function mutation at the AGL1 locus can
 be combined with any such mutation at the AGL5 locus to generate an agl1
 agl5 double mutant of the invention. Production of an exemplary agl1 agl5
 double mutant in the Brassica seed plant Arabidopsis is disclosed herein
 in Example II.
 AGL1 and AGL5 are closely related genes that have diverged relatively
 recently. While not wishing to be bound by the following, some plants can
 contain only AGL1 or only AGL5, or can contain a single ancestral gene
 related to AGL1 and AGL5. In such plants, a seed plant characterized by
 delayed seed dispersal can be produced by suppressing only expression of
 AGL1, or expression of AGL5, or expression of a single ancestral gene
 related to AGL1 and AGL5. Thus, the present invention provides a
 non-naturally occurring seed plant characterized by delayed seed
 dispersal, in which AGL1 expression is suppressed. Such a non-naturally
 occurring seed plant characterized by delayed seed dispersal can be, for
 example, an agl1 single mutant. The present invention also provides a
 non-naturally occurring seed plant characterized by delayed seed
 dispersal, in which AGL5 expression is suppressed. A non-naturally
 occurring seed plant characterized by delayed seed dispersal in which AGL5
 expression is suppressed can be, for example, an agl5 single mutant.
 The present invention further provides tissues derived from non-naturally
 occurring seed plants of the invention. In one embodiment, the invention
 provides a tissue derived from a non-naturally occurring seed plant that
 has an ectopically expressed nucleic acid molecule encoding an AGL8-like
 gene product and is characterized by delayed seed dispersal. In another
 embodiment, the invention provides a tissue derived from a non-naturally
 occurring seed plant in which AGL1 expression and AGL5expression each are
 suppressed, where the seed plant is characterized by delayed seed
 dispersal.
 As used herein, the term "tissue" means an aggregate of seed plant cells
 and intercellular material organized into a structural and functional
 unit. A particular useful tissue of the invention is a tissue that can be
 vegetatively or non-vegetatively propagated such that the seed plant from
 which the tissue was derived is reproduced. A tissue of the invention can
 be, for example, a seed, leaf, root or part thereof.
 As used herein, the term "seed" means a structure formed by the maturation
 of the ovule of a seed plant following fertilization. Such seeds can be
 readily harvested from a non-naturally occurring seed plant of the
 invention characterized by delayed seed dispersal.
 A seed plant characterized by enhanced seed dispersal also can be produced
 by manipulating expression of an AGL8-like gene product or AGL1 or AGL5.
 Suppression of AGL8-like gene product expression in a seed plant, for
 example, suppression of AGL8-like gene product expression in valve tissue,
 can be used to produce a seed plant characterized by enhanced seed
 dispersal. Ectopic expression of AGL1 or AGL5, or both, in a seed plant,
 for example, premature expression of AGL1 or AGL5, also can be used to
 produce a non-naturally occurring seed plant of the invention
 characterized by enhanced seed dispersal. The skilled person understands
 that these or other strategies of manipulating AGL8, AGL1 or AGL5
 expression can be used to produce a non-naturally occurring seed plant
 characterized by enhanced seed dispersal.
 The invention also provides a substantially purified dehiscence
 zone-selective regulatory element, which includes a nucleotide sequence
 that confers selective expression upon an operatively linked nucleic acid
 molecule in the valve margin or dehiscence zone of a seed plant, provided
 that the dehiscence zone-selective regulatory element does not have a
 nucleotide sequence consisting of nucleotides 1889 to 2703 of SEQ ID NO:4.
 As used herein, the term "dehiscence zone-selective regulatory element"
 refers to a nucleotide sequence that, when operatively linked to a nucleic
 acid molecule, confers selective expression upon the operatively linked
 nucleic acid molecule in a limited number of plant tissues, including the
 valve margin or dehiscence zone. As discussed above, the valve margin is
 the future site of the dehiscence zone and encompasses the margins of the
 outer replum as well as valve cells adjacent to the outer replum. The
 dehiscence zone, which develops in the region of the valve margin, refers
 to the group of cells that separate during the process of dehiscence,
 allowing valves to come apart from the replum and the enclosed seeds to be
 released. Thus, a dehiscence zone-selective regulatory element, as defined
 herein, confers selective expression in the mature dehiscence zone, or
 confers selective expression in the valve margin, which marks the future
 site of the dehiscence zone.
 A dehiscence zone-selective regulatory element can confer specific
 expression exclusively in cells of the valve margin or dehiscence zone or
 can confer selective expression in a limited number of plant cell types
 including cells of the valve margin or dehiscence zone. An AGL5 regulatory
 element, for example, which confers selective expression in ovules and
 placenta as well as in the dehiscence zone, is a dehiscence zone-selective
 regulatory element as defined herein. A dehiscence zone-selective
 regulatory element generally is distinguished from other regulatory
 elements by conferring selective expression in the valve margin or
 dehiscence zone without conferring expression throughout the adjacent
 carpel valves.
 The Arabidopsis AGL1 gene (SEQ ID NO:3) is shown in FIG. 7, with the
 intron-exon boundaries indicated. The Arabidopsis AGL5 gene (SEQ ID NO:4)
 is shown in FIG. 8, with the intron-exon boundaries indicated. An AGL1 or
 AGL5 regulatory element, such as a 5' regulatory element or intronic
 regulatory element, can confer selective expression in the valve margin or
 dehiscence zone and, thus, is a dehiscence-zone selective regulatory
 element as defined herein. The AGL5 gene, for example, is selectively
 expressed in the dehiscence zone, placenta and ovules, and an AGL5
 regulatory element can confer selective expression in the dehiscence zone,
 placenta and ovules upon an operatively linked nucleic acid molecule.
 The invention provides a dehiscence zone-selective regulatory element that
 is an AGL1 or AGL5 regulatory element. Such a dehiscence zone-selective
 regulatory element can be, for example, an AGL1 regulatory element. An
 AGL1 regulatory element can have, for example, the nucleotide sequence of
 a non-coding portion of the Arabidopsis AGL1 genomic sequence identified
 as SEQ ID NO:3. A dehiscence zone-selective regulatory element also can
 be, for example, an AGL5 regulatory element. An AGL5 regulatory element
 can have, for example, the nucleotide sequence of a non-coding portion of
 the Arabidopsis AGL5 genomic sequence identified as SEQ ID NO:4, provided
 that the regulatory element does not have a nucleotide sequence consisting
 of nucleotides 1889 to 2703 of SEQ ID NO:4.
 As used herein, the term "substantially the nucleotide sequence," when used
 in reference to an AGL1 or AGL5 regulatory element, means a nucleotide
 sequence having an identical sequence, or a nucleotide sequence having a
 similar, non-identical sequence that is considered to be a functionally
 equivalent sequence by those skilled in the art. For example, a dehiscence
 zone-selective regulatory element that is an AGL1 regulatory element can
 have, for example, a nucleotide sequence identical to the sequence of the
 Arabidopsis AGL1 regulatory element having nucleotides 1 to 2599 of SEQ ID
 NO:3 shown in FIG. 7, or a similar, non-identical sequence that is
 functionally equivalent. A dehiscence zone-selective regulatory element
 can have, for example, one or more modifications such as nucleotide
 additions, deletions or substitutions relative to the nucleotide sequence
 shown in FIG. 8, provided that the modified nucleotide sequence retains
 substantially the ability to confer selective expression in the valve
 margin or dehiscence zone upon an operatively linked nucleic acid
 molecule.
 It is understood that limited modifications can be made without destroying
 the biological function of an AGL1 or AGL5 regulatory element and that
 such limited modifications can result in dehiscence zone-selective
 regulatory elements that have substantially equivalent or enhanced
 function as compared to a wild type AGL1 or AGL5 regulatory element. These
 modifications can be deliberate, as through site-directed mutagenesis, or
 can be accident al such as through mutation in hosts harboring the
 regulatory element. All such modified nucleotide sequences are included in
 the definition of a dehiscence zone-selective regulatory element as long
 as the ability to confer selective express ion in the valve margin or
 dehiscence zone is substantially retained.
 A dehiscence zone-selective regulatory element can be derived from a gene
 that is an ortholog of Arabidopsis AGL1 or AGL5 and is selectively
 expressed in the valve margin or dehiscence zone of a seed plant. A
 dehiscence zone-selective regulatory element can be derived, for example,
 from an AGL1 or AGL5 ortholog of the Brassicaceae, such as a Brassica
 napus, Brassica oleracea, Brassica campestris, Brassica juncea, Brassica
 nigra or Brassica carinata AGL1 or AGL5 ortholog. A dehiscence
 zone-selective regulatory element can be derived, for example, from an
 AGL1 or AGL5 canola ortholog. A dehiscence zone-selective regulatory
 element also can be derived, for example, from a leguminous AGL1 or AGL5
 ortholog, such as a soybean, pea, chickpea, moth bean, broad bean, kidney
 bean, lima bean, lentil, cowpea, dry bean, peanut, alfalfa, lucerne,
 birdsfoot trefoil, clover, stylosanthes, lotononis bainessii, or sainfoin
 AGL1 or AGL5 ortholog.
 Dehiscence zone-selective regulatory elements also can be derived from a
 variety of other genes that are selectively expressed in the valve margin
 or dehiscence zone of a seed plant. For example, the rapeseed gene RDPG1
 is selectively expressed in the dehiscence zone (Petersen et al., Plant
 Mol. Biol. 31:517-527 (1996), which is incorporated herein by reference).
 Thus, the RDPG1 promoter or an active fragment thereof can be a dehiscence
 zone-selective regulatory element as defined herein. Additional genes such
 as the rapeseed gene SAC51 also are known to be selectively expressed in
 the dehiscence zone; the SAC51 promoter or an active fragment thereof also
 can be a dehiscence zone-selective regulatory element of the invention
 (Coupe et al., Plant Mol. Biol. 23:1223-1232 (1993), which is incorporated
 herein by reference). Further, genes selectively expressed in the
 dehiscence zone include the gene that confers selective GUS expression in
 the Arabidopsis transposant line GT140 (Sundaresan et al., Genes Devel.
 9:1797-1810 (1995), which is incorporated herein by reference). The
 skilled artisan understands that a regulatory element of any such gene
 selectively expressed in cells of the valve margin or dehiscence zone can
 be a dehiscence zone-selective regulatory element as defined herein.
 Additional dehiscence zone-selective regulatory elements can be identified
 and isolated using routine methodology. Differential screening strategies
 using, for example, RNA prepared from the dehiscence zone and RNA prepared
 from adjacent pod material can be used to isolate cDNAs selectively
 expressed in cells of the dehiscence zone (Coupe et al., supra, 1993);
 subsequently, the corresponding genes are isolated using the cDNA sequence
 as a probe.
 Enhancer trap or gene trap strategies also can be used to identify and
 isolate a dehiscence zone-selective regulatory element of the invention
 (Sundaresan et al., supra, 1995; Koncz et al., Proc. Natl. Acad. Sci. USA
 86:8467-8471 (1989); Kertbundit et al., Proc. Natl. Acad. Sci. USA
 88:5212-5216 (1991); Topping et al., Development 112:1009-1019 (1991),
 each of which is incorporated herein by reference). Enhancer trap elements
 include a reporter gene such as GUS with a weak or minimal promoter, while
 gene trap elements lack a promoter sequence, relying on transcription from
 a flanking chromosomal gene for reporter gene expression. Transposable
 elements included in the constructs mediate fusions to endogenous loci;
 constructs selectively expressed in the valve margin or dehiscence zone
 are identified by their pattern of expression. With the inserted element
 as a tag, the flanking dehiscence zone-selective regulatory element is
 cloned using, for example, inverse polymerase chain reaction methodology
 (see, for example, Aarts et al., Nature 363:715-717 (1993); see, also,
 Ochman et al., "Amplification of Flanking Sequences by Inverse PCP," in
 Innis et al., supra, 1990). The Ac/Ds transposition system of Sundaresan
 et al., supra, 1995, can be particularly useful in identifying and
 isolating a dehiscence zone-selective regulatory element of the invention.
 Dehiscence zone-selective regulatory elements also can be isolated by
 inserting a library of random genomic DNA fragments in front of a
 promoterless reporter gene and screening transgenic seed plants
 transformed with the library for dehiscence zone-selective reporter gene
 expression. The promoterless vector pROA97, which contains the npt gene
 and the GUS gene each under the control of the minimal 35S promoter, can
 be useful for such screening. The genomic library can be, for example,
 Sau3A fragments of Arabidopsis thaliana genomic DNA or genomic DNA from,
 for example, another Brassicaceae of interest (Ott et al., Mol. Gen.
 Genet. 223:169-179 (1990); Claes et al., The Plant Journal 1:15-26 (1991),
 each of which is incorporated herein by reference).
 Dehiscence zone-selective expression of a regulatory element of the
 invention can be demonstrated or confirmed by routine techniques, for
 example, using a reporter gene and in situ expression analysis. The GUS
 and firefly luciferase reporters are particularly useful for in situ
 localization of plant gene expression (Jefferson et al., EMBO J. 6:3901
 (1987); Ow et al., Science 334:856 (1986), each of which is incorporated
 herein by reference), and promoterless vectors containing the GUS
 expression cassette are commercially available, for example, from Clontech
 (Palo Alto, Calif.). To identify a dehiscence zone-selective regulatory
 element of interest such as an AGL1 or AGL5 regulatory element, one or
 more nucleotide portions of the AGL1 or AGL5 gene can be generated using
 enzymatic or PCR-based methodology (Glick and Thompson, supra, 1993; Innis
 et al., supra, 1990); the resulting segments are fused to a reporter gene
 such as GUS and analyzed as described above.
 The present invention also provides a substantially purified dehiscence
 zone-selective regulatory element that confers selective expression upon
 an operatively linked nucleic acid molecule in the valve margin or
 dehiscence zone of a seed plant, where the element is an AGL1 regulatory
 element having at least fifteen contiguous nucleotides of one of the
 following nucleotide sequences: nucleotides 1 to 2599 of SEQ ID NO:3;
 nucleotides 2833 to 4128 of SEQ ID NO:3; nucleotides 4211 to 4363 of SEQ
 ID NO:3; nucleotides 4426 to 4554 of SEQ ID NO:3; nucleotides 4655 to
 4753; nucleotides 4796 to 4878 of SEQ ID NO:3; nucleotides 4921 to 5028 of
 SEQ ID NO:3; or nucleotides 5361 to 5622 of SEQ ID NO:3. A substantially
 purified dehiscence zone-selective regulatory element that is an AGL1
 regulatory element can have, for example, at least 16, 18, 20, 25, 30, 40,
 50, 100 or 500 contiguous nucleotides of one of the portions of SEQ ID
 NO:3 described above.
 The present invention also provides a substantially purified dehiscence
 zone-selective regulatory element that confers selective expression upon
 an operatively linked nucleic acid molecule in the valve margin or
 dehiscence zone of a seed plant, where the element is an AGL5 regulatory
 element having at least fifteen contiguous nucleotides of one of the
 following nucleotide sequences: nucleotides 1 to 1888 of SEQ ID NO:4;
 nucleotides 2928 to 5002 of SEQ ID NO:4; nucleotides 5085 to 5204 of SEQ
 ID NO:4; nucleotides 5367 to 5453 of SEQ ID NO:4; nucleotides 5496 to
 5602; nucleotides 5645 to 5734 of SEQ ID NO:4; or nucleotides 6062 to 6138
 of SEQ ID NO:4. A substantially purified dehiscence zone-selective
 regulatory element that is an AGL5 regulatory element can have, for
 example, at least 16, 18, 20, 25, 30, 40, 50, 100 or 500 contiguous
 nucleotides of one of the portions of SEQ ID NO:4 described above.
 A proximal fragment of the Arabidopsis AGL5 promoter has been described
 (Savidge et al., The Plant Cell 7:721-733 (1995)). However, this fragment
 (shown as nucleotides 1889 to 2703 in FIG. 8) lacks many of the distal
 regulatory elements contained in the entire Arabidopsis AGL5 genomic
 sequence disclosed herein (SEQ ID NO:4). The present invention provides
 approximately 2.7 kb of Arabidopsis AGL5 5' flanking sequence, including
 the variety of regulatory elements contained therein. The disclosed
 Arabidopsis AGL5 5' flanking sequence contains a larger complement of
 regulatory elements involved in regulating expression of the endogenous
 AGL5 gene in vivo and, therefore, can be particularly useful for
 dehiscence zone-selective expression.
 A nucleotide sequence consisting of the promoter proximal region of
 Arabidopsis AGL5 (nucleotides 1889 to 2703 of SEQ ID NO:4) is explicitly
 excluded from a dehiscence zone-selective regulatory element of the
 invention. However, a dehiscence zone-selective regulatory element can
 include nucleotides 1889 to 2703 of SEQ ID NO:4, together with one or more
 contiguous nucleotides, for example, of the nucleotide sequence shown as
 positions 1 to 1888 of SEQ ID NO:4. A dehiscence zone-selective regulatory
 element of the invention can have, for example, at least 15 contiguous
 nucleotides of SEQ ID NO:4, including at least one, two, four, six, ten,
 twenty or thirty or more contiguous nucleotides of the nucleotide sequence
 shown as positions 1 to 1888 of SEQ ID NO:4.
 In view of the definition of a dehiscence zone-selective regulatory
 element, it should be recognized, for example, that a portion of the
 Arabidopsis AGL5 gene having only the sequence shown as nucleotides 1889
 to 2703 in FIG. 8 (SEQ ID NO:4), is not a dehiscence zone-selective
 regulatory element as defined herein. However, a portion of an Arabidopsis
 AGL5 gene having nucleotides 1885 to 2703 of SEQ ID NO:4 is considered a
 dehiscence zone-selective regulatory element, provided that the element
 confers selective expression upon an operatively linked nucleic acid
 molecule in a limited number of plant tissues, including the valve margin
 or dehiscence zone. Similarly, a portion of an Arabidopsis AGL5 gene
 having a subpart of the promoter proximal region of AGL5 also can be a
 dehiscence zone-selective regulatory element as defined herein, provided
 that this subpart can confer selective expression upon an operatively
 linked nucleic acid molecule in a limited number of plant tissues,
 including the valve margin or dehiscence zone of a seed plant. Thus, for
 example, a regulatory element having the sequence of nucleotides 1889 to
 2000 can be a dehiscence zone-selective regulatory element of the
 invention, provided that this element confers selective expression upon an
 operatively linked element in the valve margin or dehiscence zone of a
 seed plant.
 The present invention also provides a recombinant nucleic acid molecule
 that includes a dehiscence zone-selective regulatory element operatively
 linked to a nucleic acid molecule encoding a cytotoxic gene product.
 Further provided herein is a non-naturally occurring seed plant of the
 invention that is characterized by delayed seed dispersal due to
 expression of a recombinant nucleic acid molecule having a dehiscence
 zone-selective regulatory element operatively linked to a nucleic acid
 molecule encoding a cytotoxic gene product.
 A cytotoxic gene product is a gene product that causes the death of the
 cell in which it is expressed and, preferably, does not result in the
 death of cells other than the cell in which it is expressed. Thus,
 expression of a cytotoxic gene product from a dehiscence zone-selective
 regulatory element can be used to ablate the dehiscence zone without
 disturbing neighboring cells of the replum or valve. A variety of
 cytotoxic gene products useful in seed plants are known in the art
 including, for example, diphtheria toxin A chain polypeptides; RNase T1;
 Barnase RNase; ricin toxin A chain polypeptides; and herpes simplex virus
 thymidine kinase (tk) gene products. While the diphtheria toxin A chain,
 RNase T1 and Barnase RNase are preferred cytotoxic gene products, the
 skilled person recognizes that these, or other cytotoxic gene products can
 be used with a dehiscence zone-selective regulatory element to generate a
 non-naturally occurring seed plant characterized by delayed seed
 dispersal.
 Diphtheria toxin is the naturally occurring toxin of Cornebacterium
 diphtheriae, which catalyzes the ADP-ribosylation of elongation factor 2,
 resulting in inhibition of protein synthesis and consequent cell death
 (Collier, Bacteriol. Rev. 39:54-85 (1975)). A single molecule of the fully
 active toxin is sufficient to kill a cell (Yamaizumi et al., Cell
 15:245-250 (1978)). Diphtheria toxin has two subunits: the diphtheria
 toxin B chain directs internalization to most eukaryotic cells through a
 specific membrane receptor, whereas the A chain encodes the toxic
 catalytic domain. The catalytic DT-A chain does not include a signal
 peptide and is not secreted. Further, any DT-A released from dead cells in
 the absence of the diphtheria toxin B chain is precluded from cell
 attachment. Thus, DT-A is cell autonomous and directs killing only of the
 cells in which it is expressed without apparent damage to neighboring
 cells. The DT-A expression cassette of Palmiter et al., which contains the
 193 residues of the A chain engineered with a synthetic ATG and lacking
 the native leader sequence, is particularly useful in the seed plants of
 the invention (Palmiter et al., Cell 50:435-443 (1987); Greenfield et al.,
 Proc. Natl. Acad. Sci., USA 80:6853-6857 (1983), each of which is
 incorporated herein by reference)
 RNase T1 of Aspergillus oryzae and Barnase RNase of Bacillus
 amylolique-faciens also are cytotoxic gene products useful in the seed
 plants of the invention (Thorsness and Nasrallah, Methods in Cell Biology
 50:439-448 (1995)). Barnase RNase may be more generally toxic to plants
 than RNase T1 and, thus, is preferred in the methods of the invention.
 Ricin, a ribosome-inactivating protein produced by castor bean seeds, also
 is a cytotoxic gene product useful in a non-naturally occurring seed plant
 of the invention. The ricin toxin A chain polypeptide can be used to
 direct cell-specific ablation as described, for example, in Moffat et al.,
 Development 114:681-687 (1992). Plant ribosomes are variably susceptible
 to the plant-derived ricin toxin. The skilled person understands that the
 toxicity of ricin depends is variable and should be assessed for toxicity
 in the seed plant species of interest (see Olsnes and Pihl, Molecular
 Action of Toxins and Viruses, pages 51-105, Amsterdam: Elsevier Biomedical
 Press (1982)).
 Further provided herein is a plant expression vector including a dehiscence
 zone-selective regulatory element. A plant expression vector can include,
 if desired, a nucleic acid molecule encoding an AGL8-like gene product in
 addition to the dehiscence zone-selective regulatory element.
 The term "plant expression vector," as used herein, is a self-replicating
 nucleic acid molecule that provides a means to transfer an exogenous
 nucleic acid molecule into a seed plant host cell and to express the
 molecule therein. Plant expression vectors encompass vectors suitable for
 Agrobacterium-mediated transformation, including binary and cointegrating
 vectors, as well as vectors for physical transformation.
 Plant expression vectors can be used for transient expression of the
 exogenous nucleic acid molecule, or can integrate and stably express the
 exogenous sequence. One skilled in the art understands that a plant
 expression vector can contain all the functions needed for transfer and
 expression of an exogenous nucleic acid molecule; alternatively, one or
 more functions can be supplied in trans as in a binary vector system for
 Agrobacterium-mediated transformation.
 In addition to a dehiscence zone-selective regulatory element, a plant
 expression vector of the invention can contain, if desired, additional
 elements. A binary vector for Agrobacterium-mediated transformation
 contains one or both T-DNA border repeats and can also contain, for
 example, one or more of the following: a broad host range replicon, an ori
 T for efficient transfer from E. coli to Agrobacterium, a bacterial
 selectable marker such as ampicillin and a polylinker containing multiple
 cloning sites.
 A plant expression vector for physical transformation can have, if desired,
 a plant selectable marker in addition to a dehiscence zone-selective
 regulatory element in vectors such as pBR322, pUC, PGEM and M13, which are
 commercially available, for example, from Pharmacia (Piscataway, N.J.) or
 Promega (Madison, Wis.). In plant expression vectors for physical
 transformation of a seed plant, the T-DNA borders or the ori T region can
 optionally be included but provide no advantage.
 The present invention also provides a kit for producing a transgenic seed
 plant characterized by delayed seed dispersal. A kit of the invention
 contains a dehiscence zone-selective regulatory element. If desired, the
 dehiscence zone-selective regulatory element can be operatively linked to
 a nucleic acid molecule encoding an AGL8-like gene product.
 The following examples are intended to illustrate but not limit the present
 invention.
 EXAMPLE I
 PRODUCTION OF A 35S-AGL8 TRANSGENIC ARABIDOPSIS PLANT DISPLAYING A COMPLETE
 LACK OF DEHISCENCE
 This example describes methods for producing a transgenic Arabidopsis plant
 lacking normal dehiscence due to constitutive AGL8 expression.
 Full-length AGL8 was prepared by polymerase chain reaction amplification
 using primer AGL8 5-.gamma. (SEQ ID NO:9;
 5'-CCGTCGACGATGGGAAGAGGTAGGGTT-3') and primer OAM14 (SEQ ID NO:10;
 5'-AATCATTACCAAGATATGAA-3'), and subsequently cloned into the SalI and
 BamHI sites of expression vector pBIN-JIT, which was modified from pBIN19
 to include the tandem CaMV 35S promoter, a polycloning site and the CaMV
 polyA signal. Arabidopsis was transformed using the in planta method of
 Agrobacterium-mediated transformation essentially as described in Bechtold
 et al., C.R. Acad. Sci. Paris 316:1194-1199 (1993), which is incorporated
 herein by reference. Kanamycin-resistant lines were analyzed for the
 presence of the 35S-AGL8 construct by PCR using a primer specific for the
 35S promoter and a primer specific for the AGL8 cDNA, which produced two
 fragments of 850 and 550 bp in the 35S-AGL8 transgenic plants. These
 fragments were absent in plants that had not been transformed with the
 35S-AGL8 construct.
 The phenotype of approximately 35 35S::AGL8 lines was analyzed. Of the 35
 lines, 7 lines exhibited a complete lack of dehiscence. In these lines,
 the mature fruits did not release their seeds unless opened manually.
 Several of the remaining 35S::AGL8 lines exhibited delayed dehiscence,
 whereby seeds were released at least a week later than in wild type
 Arabidopsis plants.
 EXAMPLE II
 PRODUCTION OF AN ARABIDOPSIS agl1 agl5 double mutant DISPLAYING A COMPLETE
 LACK OF DEHISCENCE
 This example describes the production of an agl1 agl5 double mutant
 displaying a complete lack of normal dehiscence.
 A. Production of an agl5 Mutant by Homologous Recombination
 A PCR-based assay of transgenic plants was used to identify targeted
 insertions into AGL5 as described in Kempin et al., Nature 389:802-803
 (1997), which is incorporated herein by reference. The targeting construct
 consisted of a kanamycin-resistance cassette that was inserted between
 approximately 3 kb and 2 kb segments representing the 5' and 3' regions of
 the AGL5 gene, respectively. A successfully targeted insertion produces a
 1.6 kb deletion within the AGL5 gene such that the targeted allele encodes
 only the first 42 of 246 amino acid residues, and only 26 of the 56 amino
 acids comprising the DNA-binding MADS-domain. The recombination event also
 results in the insertion of the 2.5 kb kanamycin-resistance cassette
 within the AGL5 coding sequence.
 750 kanamycin-resistant transgenic lines were produced by
 Agrobacterium-mediated transformation, and pools of transformants were
 analyzed using a PCR assay as described below to determine if any of these
 primary transformants had generated the desired targeted insertion into
 AGL5. A single line was identified that appeared to contain the
 anticipated insertion, and this line was allowed to self-pollinate to
 permit further analyses in subsequent generations. Genomic DNA from the
 homozygous mutant plants was analyzed with more than four different
 restriction enzymes and by several distinct PCR amplifications, and all
 data were consistent with the desired targeting event. The regions
 flanking the AGL5 gene also were analyzed to verify that there were no
 detectable deletions or rearrangements of sequences outside of AGL5.
 The kanamycin-resistance cassette within the AGL5 targeting construct
 contains sequences that specify transcription termination such that little
 or no AGL5 RNA was expected in the homozygous mutant plants. Using a probe
 specific for the 3' portion of the AGL5 cDNA, AGL5 transcripts were
 detected in wild-type but not in agl5 mutant plants. These data indicate
 that the targeted disruption of the AGL5 gene represents a
 loss-of-function allele.
 Characterization of the agl5 line indicated that the phenotype of this
 transgenic was not different from wild type Arabidopsis.
 The AGL5 knockout (KO) construct was prepared in vector pZM104A, which
 carries the kanamycin-resistance cassette flanked by several cloning sites
 (Miao and Lam, Plant J. 7:359-365 (1995), which is incorporated herein by
 reference). Vector pZM104A also contains the gene encoding
 .delta.-glucuronidase (GUS), which allows the differentiation of
 non-homologous from homologous integration events. The 3 kb region
 representing the 5' portion of AGL5 was obtained by PCR amplification
 using primer SEQ ID NO:11 (5'-CGGATAGCTCGAATATCG-3') and primer SEQ ID
 NO:12 (5'-AACCATTGCGTCGTTTGC-3'). The resulting fragment was cloned into
 vector pCRII (Invitrogen), and an EcoRI fragment excised and inserted into
 the EcoRI site of pZM104A. The 3' portion of AGL5 was excised as an XbaI
 fragment from an AGL5 genomic clone in the vector pCIT30 (Ma et al., Gene
 117:161-167 (1992), which is incorporated by reference herein) and
 inserted into the XbaI site of pZM104A. The resulting plasmid, designated
 AGL5 KO, was used in Agrobacterium-mediated infiltration of wild-type
 Arabidopsis plants of the Columbia ecotype. The knockout construct was
 derived from Landsberg erecta genomic DNA.
 Plants containing a homologous recombination event at the AGL5 genomic
 locus were identified as follows. Approximately 750 primary (Ti)
 kanamycin-resistant transformants were selected, and DNA was extracted
 from individual leaves in pools representing ten plants as described in
 Edwards et al., Nucleic Acids Research 19:1349 (1991), which is
 incorporated by reference herein. To identify a pool that contained a
 candidate targeted disruption, isolated DNAs were subjected to PCR
 amplification using primer SEQ ID NO:13 (5'-GTAATTACCAGGCAAGGACTCTCC-3'),
 which represents AGL5 genomic sequence that is not contained within the
 AGL5 KO construct, and primer SEQ ID NO:14
 (5'-GTCATCGGCGGGGGTCATAACGTG-3'), which is specific for the
 kanamycin-resistance cassette. Amplified products were size fractionated
 on agarose gels, and used for standard DNA blotting assays with probe 1.
 One pool of ten plants revealed the anticipated hybridizing band of the
 correct size, and this pool was subsequently broken down into individual
 plants. A single (T1) plant was identified that appeared to contain the
 desired event, and this plant was allowed to self-pollinate for analyses
 in subsequent generations. This T1 plant was shown to contain the
 GUS-reporter gene, indicating that in addition to the putative homologous
 integration event, there were independent non-homologous events.
 Segregation in the subsequent generations allowed the identification of
 plants that no longer contained the GUS-reporter gene, and it was these
 lines that were used for subsequent analyses.
 Plants homozygous for the disruption were identified by PCR amplification
 using primers SEQ ID NO:15 (5'-GAGGATAGAGAACACTACGAATCG-3') and SEQ ID
 NO:16 (5'-CAGGTCAAGTCAATAGATTC-3'), which yielded a single 1.5 kb product
 in wild type plants, and a single 2.6 kb product in the mutant. Further
 confirmation that these plants contained the desired disruption was
 obtained by PCR amplification with primers SEQ ID NO:17
 (5'-CAGAATTTAGTGAATAATATTG-3') and SEQ ID NO:14, which gave the expected
 amplified product in the mutant but no product in wild-type plants.
 To confirm that the desired disruption had occurred, a series of genomic
 DNA blots representing wild-type and homozygous mutant (T4 generation)
 plants were analyzed. Probe 1 hybridized to the expected 3.9 kb XbaI
 fragment in wild-type and mutant plants, whereas the 1.3 kb XbaI fragment
 was present only in wild-type. This same probe hybridized to a 6 kb EcoRI
 fragment in wild-type and to the expected 4.1 and 2.8 kb EcoRI fragments
 in the mutant. Additional digests with BglII and with HindIII confirmed
 that the mutant plants contained the desired targeted event. To confirm
 that there were no detectable deletions or rearrangements outside the
 targeted region, genomic DNA blots of wild type and homozygous mutant
 plants were further analyzed. Probe 2 hybridized in wild-type and mutant
 DNAs to the expected 2.9 kb XmnI fragment, the 1.5 kb and 0.4 kb HincII
 fragments, and the 0.6 kb HindIII fragment. Probe 3 hybridized in
 wild-type and mutant DNAs to the 9 kb ScaI fragment, the 3.9 kb XbaI
 fragment, and the 1.8 kb NdeI fragments. The faintly-hybridizing bands in
 the ScaI digests represent fragments that span the insertion site, and
 are, as expected, different sizes in wild-type and agl5 mutant plants.
 RNA blotting analyses were performed as follows. Approximately 6 .mu.g of
 polyA+ RNA was purified using Dynabeads (Dynal) from wild-type and agl5
 mutant inflorescences, size fractionated and hybridized using standard
 procedures (Crawford et al., Proc. Natl. Acad. Sci. USA 83:8073-8076
 (1986), which is incorporated herein by reference) using a gel-purified
 450 bp HindIII-EcoRI fragment from pCIT2242 (Ma et al., supra, 1991)
 specific for the 3' end of the AGL5 cDNA. The same filter was subsequently
 stripped and re-hybridized with a tubulin-specific probe (Marks et al.,
 Plant Mol. Biol. 10:91-104 (1987), which is incorporated herein by
 reference). Hybridization with the tubulin probe verified that
 approximately equal amounts of RNA were present in each lane.
 B. Production of an agl1 Mutant
 A PCR-based screen was used to identify a T-DNA insertion into the AGL1
 gene essentially as described in Krysan et al., supra, 1996.
 RNA blotting analyses demonstrated that AGL1 RNA was not expressed. The
 agl1 mutant displayed essentially a wild type phenotype.
 C. Production and Characterization of an agl1 agl5 Double Mutant
 agl1 agl5 double mutants were generated by crossing the agl1 and agl5
 single mutants. RNA blotting experiments of the agl1 agl5 double mutant
 are performed as described above. The results indicate that neither AGL1
 nor AGL5 RNA is expressed in the agl1 agl5 double mutant.
 In contrast to the agl1 and agl5 single mutants, which had essentially the
 phenotype of wild type Arabidopsis, analyses of the agl1 agl5 double
 mutant by scanning electron microscopy indicated that the dehiscence zone
 failed to develop normally. Furthermore, the mature fruits of the agl1
 agl5 double mutant failed to dehisce. This delayed seed dispersal
 phenotype was similar to AGL8 gain-of-function phenotype seen in 35S-AGL8
 transgenic plants. These results indicate that the AGL1 and AGL5 genes are
 functionally redundant and that their encoded gene products regulate pod
 dehiscence. The similarity of the 35S::AGL8 and agl1 agl5 double mutant
 phenotypes, as well the yeast two-hybrid results described below, indicate
 that AGL1 and AGL8 or AGL5 and AGL8 can interact to regulate the
 dehiscence process.
 D. Analysis of Dehiscence Phenotypes Under Various Conditions
 Studies of pod dehiscence in Brassica napus L. using transmission electron
 microscopic analyses have shown that the middle lamella of the dehiscence
 zone cells degenerates during dehiscence, allowing the valves to separate
 from the replum (Petersen et al., supra, 1996). Similar analyses are
 performed on the agl1 agl5 double mutant as well as wild type Arabidopsis
 and agl1 and agl5 single mutants.
 Previous studies have shown that pod dehiscence is greater when
 temperatures are high and the relative humidity is low. The dehiscence
 phenotype of the agl1 agl5 double mutant described above was observed for
 plants grown under continuous-light at 25 degrees C. In order to determine
 if the phenotype of agl1 agl5 double mutants is sensitive to environmental
 conditions, the analyses described above are repeated under various
 environmental conditions including varying temperature, varying humidity
 and short-day versus continuous light conditions.
 EXAMPLE III
 PRODUCTION OF A TRANSGENIC ARABIDOPSIS PLANT EXPRESSING AGL8 UNDER CONTROL
 OF THE AGL1 PROMOTER
 This example demonstrates that a transgenic seed plant expressing AGL8
 under control of a dehiscence zone-selective promoter is characterized by
 delayed seed dispersal.
 AGL1::AGL8 Transgenic Plants
 Ectopic expression of AGL8 under control of the 35S promoter prevents pod
 shatter since the dehiscence zone fails to differentiate normally.
 However, constitutive AGL8 expression conferred by the 35S promoter also
 results in other changes, including early flowering. In order to
 specifically control dehiscence, AGL8 is expressed from a dehiscence
 zone-selective regulatory element, such as one derived from a regulated
 promoter that is normally expressed in valve margin, as described below.
 An AGL8 expression construct under control of the dehiscence zone-selective
 2.5 kb AGL1 promoter fragment and first AGL1 intronic sequence is prepared
 as follows. The 2.5 kb AGL1 promoter fragment is amplified by PCR with
 primers AGL1pds (SEQ ID NO:18; 5'-GCCAGAGATAATGCTATTCC-3') and AGL1pus
 (SEQ ID NO:19; 5'-CATTGATCCATATATGACATCAC-3'), and the first coding exon
 of AGL8 is amplified with oligos AGL8eds (SEQ ID NO:20;
 5'-GTGATGTCATATATGGATCAATGGGAAGAGGTAGGGTTCAG-3') and AGL8eus (SEQ ID
 NO:21; 5'-CAAGAGTCGGTGGAATATTCG-3'). In addition, the first intron of
 AGL1, which can contain regulatory elements, is amplified with oligos
 AGL1ids (SEQ ID NO:22; 5'-CGAATATTCCACCGACTCTTGGTACGCTTC TCCTACTCTAT-3')
 and AGL1iup (SEQ ID NO:23; 5'-CTAATAAGTAAGATCGCGGAA-3'). The remainder of
 the AGL8 coding region is amplified with oligos AGL8rds (SEQ ID NO:24;
 5'-TTCCGCGATCTTACTTATTAGCATGGAGAGGATACTTGAAC-3') and OAM14 (SEQ ID NO:10).
 Using PCR with oligos AGL1pds (SEQ ID NO:18) and OAM14 (SEQ ID NO:10), the
 four fragments are combined in the following order: AGL1 promoter, first
 AGL8 exon, first AGL1 intron and remainder of AGL8 coding sequence. The
 resulting 4.6 kb fragment is cloned into vector pCFM83, which is a vector
 based on pBIN19 that is modified to contain a BASTA resistance gene and 3'
 NOS termination sequence.
 A second AGL8 expression construct, in which AGL8 is under control of the
 dehiscence zone-selective 2.5 kb AGL1 promoter fragment alone, is prepared
 as follows. The 2.5 kb AGL1 promoter fragment is amplified by PCR with
 oligo AGL1pds (SEQ ID NO:18) and AGL1pus (SEQ ID NO:19), and the coding
 region of AGL8 amplified with oligos AGL8eds (SEQ ID NO:20) and OAM14 (SEQ
 ID NO:10). Using PCR with oligos AGL1 pds (SEQ ID NO:18) and OAM14 (SEQ ID
 NO:10), the 3.5 kb fragment is cloned into vector pCFM83.
 Arabidopsis plants are transformed with the two AGL1-AGL8 constructs
 described above. BASTA resistant plants containing the AGL1::AGL8
 transgene with or without the AGL1 intron are selected. Phenotypic
 analysis indicates that transformed plants containing either of these
 constructs are characterized by delayed dehiscence. However, the
 AGL1::AGL8 transgenic plants differ from 35S::AGL8 transgenic plants in
 that an enlarged fruit or early flowering phenotype generally is not seen.
 These results indicate that a transgenic seed plant expressing AGL8 under
 control of an AGL1 dehiscence zone-selective regulatory element is
 characterized by delayed seed dispersal.
 EXAMPLE IV
 AGL8 INTERACTS WITH AGL5 IN YEAST
 This example demonstrates that, in a yeast two-hybrid system, the AGL8 gene
 product interacts with AGL5.
 The "interaction trap" of Finley and Brent (Gene Probes: A Practical
 Approach (1994); see, also Gyuris et al., Cell 75:791-803 (1993)) is a
 variation of the yeast two-hybrid system of Fields and Song, Nature
 340:245-246 (1989). In this system, a first protein is fused to a
 DNA-binding domain, and a second is fused to a transcriptional activation
 domain. An interaction between the Arabidopsis AGL5 and AGL8 gene products
 was assayed by activation of a lacZ reporter gene.
 The "bait" and "prey" constructs were prepared in single copy centromere
 plasmids pBI-880 and pBI-771, respectively, which each contain the
 constitutive ADH1 promoter and are essentially as described by Chevray and
 Nathans, Proc. Natl. Acad. Sci. USA 89:5789-5793 (1992). The bait
 construct contains the GAL4 DNA-binding domain (amino acids 1 to 147)
 fused to the full-length AGL8 coding sequence. The prey construct has the
 full-length coding sequence of AGL5 fused to the GAL4 transcriptional
 activation domain (amino acids 768-881), following a nuclear localization
 sequence. The bait and prey constructs were assayed in the YPB2 strain of
 S. cerevisiae, which is deficient for GAL4 and GAL80 and which contains an
 integrated lacZ reporter gene under control of GAL1 promoter elements
 (Feilotter et al., Nucleic Acids Research 22:1502-1503 (1994)).
 An interaction of the AGL8 "bait" and AGL5 "prey" was demonstrated in the
 YPB2 strain by the development of blue colonies on X-GAL containing media.
 Control "bait"-"prey" combinations, including the GAL4(1-147) DNA binding
 domain and GAL4 transcriptional activation domain only produced only white
 colonies. These results demonstrate that AGL8 can interact with AGL5 in
 yeast and indicate that the AGL8 and AGL5 plant MADS box gene products
 also can interact in seed plants.
 All journal article, reference, and patent citations provided above, in
 parentheses or otherwise, whether previously stated or not, are
 incorporated herein by reference.
 Although the invention has been described with reference to the examples
 above, it should be understood that various modifications can be made
 without departing from the spirit of the invention. Accordingly, the
 invention is limited only by the following claims.
 SEQUENCE LISTING
 (1) GENERAL INFORMATION:
 (iii) NUMBER OF SEQUENCES: 24
 (2) INFORMATION FOR SEQ ID NO: 1:
 (i) SEQUENCE CHARACTERISTICS:
 (A) LENGTH: 1062 base pairs
 (B) TYPE: nucleic acid
 (C) STRANDEDNESS: single
 (D) TOPOLOGY: linear
 (ii) MOLECULE TYPE: cDNA
 (vi) ORIGINAL SOURCE:
 (A) ORGANISM: not provided
 (ix) FEATURE:
 (A) NAME/KEY: CDS
 (B) LOCATION: 101..827
 (ix) FEATURE:
 (A) NAME/KEY: misc_feature
 (B) LOCATION: 1062
 (D) OTHER INFORMATION: /note= "There is a poly(A) tail at
 the end."
 (ix) FEATURE:
 (A) NAME/KEY: misc_feature
 (B) LOCATION: 1..1062
 (D) OTHER INFORMATION: /note= "Nucleotide and Deduced
 Amino Acid Sequences of the AGL8 cDNA clone."
 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:1
 CCCAGAGAGA CATAAGAAAG AAAGAGAGAG AGAGATACTT TGGTCATTTC AGGGTTGTCG 60
 TTTCTCTCTC TTGTTCTTGA GATTTTGAAG AGAGAGAGAT ATG GGA AGA GGT AGG 115
 Met Gly Arg Gly Arg
 1 5
 GTT CAG CTG AAG AGG ATA GAG AAC AAG ATC AAT AGG CAA GTT ACT TTC 163
 Val Gln Leu Lys Arg Ile Glu Asn Lys Ile Asn Arg Gln Val Thr Phe
 10 15 20
 TCA AAG AGA AGG TCT GGT TTG CTC AAG AAA GCT CAT GAG ATC TCT GTT 211
 Ser Lys Arg Arg Ser Gly Leu Leu Lys Lys Ala His Glu Ile Ser Val
 25 30 35
 CTC TGC GAT GCT GAG GTT GCT CTC ATC GTC TTC TCT TCC AAA GGC AAA 259
 Leu Cys Asp Ala Glu Val Ala Leu Ile Val Phe Ser Ser Lys Gly Lys
 40 45 50
 CTC TTC GAA TAT TCC ACC GAC TCT TGC ATG GAG AGG ATA CTT GAA CGC 307
 Leu Phe Glu Tyr Ser Thr Asp Ser Cys Met Glu Arg Ile Leu Glu Arg
 55 60 65
 TAT GAT CGC TAT TTA TAT TCA GAC AAA CAA CTT GTT GGC CGA GAC GTT 355
 Tyr Asp Arg Tyr Leu Tyr Ser Asp Lys Gln Leu Val Gly Arg Asp Val
 70 75 80 85
 TCA CAA AGT GAA AAT TGG GTT CTA GAA CAT GCT AAG CTC AAG GCA AGA 403
 Ser Gln Ser Glu Asn Trp Val Leu Glu His Ala Lys Leu Lys Ala Arg
 90 95 100
 GTT GAG GTA CTT GAG AAG AAC AAA AGG AAT TTT ATG GGG GAA GAT CTT 451
 Val Glu Val Leu Glu Lys Asn Lys Arg Asn Phe Met Gly Glu Asp Leu
 105 110 115
 GAT TCG TTG AGC TTG AAG GAG CTC CAA AGC TTG GAG CAT CAG CTC GAT 499
 Asp Ser Leu Ser Leu Lys Glu Leu Gln Ser Leu Glu His Gln Leu Asp
 120 125 130
 GCA GCT ATC AAG AGC ATT AGG TCA AGA AAG AAC CAA GCT ATG TTC GAA 547
 Ala Ala Ile Lys Ser Ile Arg Ser Arg Lys Asn Gln Ala Met Phe Glu
 135 140 145
 TCC ATA TCT GCG CTC CAG AAG AAG GAT AAA GCC TTG CAA GAT CAC AAC 595
 Ser Ile Ser Ala Leu Gln Lys Lys Asp Lys Ala Leu Gln Asp His Asn
 150 155 160 165
 AAT TCG CTT CTC AAA AAG ATT AAG GAG AGG GAG AAG AAA ACG GGT CAG 643
 Asn Ser Leu Leu Lys Lys Ile Lys Glu Arg Glu Lys Lys Thr Gly Gln
 170 175 180
 CAA GAA GGA CAA TTA GTC CAA TGC TCC AAC TCT TCT TCA GTT CTT CTG 691
 Gln Glu Gly Gln Leu Val Gln Cys Ser Asn Ser Ser Ser Val Leu Leu
 185 190 195
 CCT CAA TAC TGC GTA ACC TCC TCC AGA GAT GGC TTT GTG GAG AGA GTT 739
 Pro Gln Tyr Cys Val Thr Ser Ser Arg Asp Gly Phe Val Glu Arg Val
 200 205 210
 GGG GGA GAG AAC GGT GGT GCA TCG TCG TTG ACG GAA CCA AAC TCT CTG 787
 Gly Gly Glu Asn Gly Gly Ala Ser Ser Leu Thr Glu Pro Asn Ser Leu
 215 220 225
 CTT CCG GCT TGG ATG TTA CGT CCT ACC ACT ACG AAC GAG T AGAACTATCT 837
 Leu Pro Ala Trp Met Leu Arg Pro Thr Thr Thr Asn Glu
 230 235 240
 CACTCTTTAT AATATAATGA TAATATAATT AATGTTTAAT ATTTTCATAA CATTCAGCAT 897
 TTTTTTGGTG ACTTATACTC ATTATTAATA CCGATATGTT TTAGCTAGTC ATATTATATG 957
 TATGATGGAA CTCCGTTGTC GAGACGTATG TACGTAAGCT ATCATTAGAT TCACTGCGTC 1017
 TTAAGAACAA AGATTCATAT CTTGGTAATG ATTTCTCATG AAATA 1062
 (2) INFORMATION FOR SEQ ID NO: 2:
 (i) SEQUENCE CHARACTERISTICS:
 (A) LENGTH: 242 amino acids
 (B) TYPE: amino acid
 (D) TOPOLOGY: linear
 (ii) MOLECULE TYPE: protein
 (vi) ORIGINAL SOURCE:
 (A) ORGANISM: not provided
 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:2
 Met Gly Arg Gly Arg Val Gln Leu Lys Arg Ile Glu Asn Lys Ile Asn
 1 5 10 15
 Arg Gln Val Thr Phe Ser Lys Arg Arg Ser Gly Leu Leu Lys Lys Ala
 20 25 30
 His Glu Ile Ser Val Leu Cys Asp Ala Glu Val Ala Leu Ile Val Phe
 35 40 45
 Ser Ser Lys Gly Lys Leu Phe Glu Tyr Ser Thr Asp Ser Cys Met Glu
 50 55 60
 Arg Ile Leu Glu Arg Tyr Asp Arg Tyr Leu Tyr Ser Asp Lys Gln Leu
 65 70 75 80
 Val Gly Arg Asp Val Ser Gln Ser Glu Asn Trp Val Leu Glu His Ala
 85 90 95
 Lys Leu Lys Ala Arg Val Glu Val Leu Glu Lys Asn Lys Arg Asn Phe
 100 105 110
 Met Gly Glu Asp Leu Asp Ser Leu Ser Leu Lys Glu Leu Gln Ser Leu
 115 120 125
 Glu His Gln Leu Asp Ala Ala Ile Lys Ser Ile Arg Ser Arg Lys Asn
 130 135 140
 Gln Ala Met Phe Glu Ser Ile Ser Ala Leu Gln Lys Lys Asp Lys Ala
 145 150 155 160
 Leu Gln Asp His Asn Asn Ser Leu Leu Lys Lys Ile Lys Glu Arg Glu
 165 170 175
 Lys Lys Thr Gly Gln Gln Glu Gly Gln Leu Val Gln Cys Ser Asn Ser
 180 185 190
 Ser Ser Val Leu Leu Pro Gln Tyr Cys Val Thr Ser Ser Arg Asp Gly
 195 200 205
 Phe Val Glu Arg Val Gly Gly Glu Asn Gly Gly Ala Ser Ser Leu Thr
 210 215 220
 Glu Pro Asn Ser Leu Leu Pro Ala Trp Met Leu Arg Pro Thr Thr Thr
 225 230 235 240
 Asn Glu
 (2) INFORMATION FOR SEQ ID NO: 3:
 (i) SEQUENCE CHARACTERISTICS:
 (A) LENGTH: 5622 base pairs
 (B) TYPE: nucleic acid
 (C) STRANDEDNESS: unknown
 (D) TOPOLOGY: unknown
 (vi) ORIGINAL SOURCE:
 (A) ORGANISM: not provided
 (ix) FEATURE:
 (A) NAME/KEY: misc_feature
 (B) LOCATION: 1..5622
 (D) OTHER INFORMATION: /label= AGL1_promoter
 /note= "Nucleotide sequence of the AGL1 promoter."
 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:3
 AGATCTGCAA CAGTGAAAAG AGAAAACAAA ATGGACTTGA AGAGGTTTTG ACAATGCCAG 60
 AGATAATGCT TATTCCCTAA TATGTTGCCA GCCAAGTGTC AAATTGGCTT TTTAAATATG 120
 GATTTCTGTA TCAGTGGTCA TATTTGTGGA TCCAACGTAT TCATCATCAA GTTCTCAAGT 180
 TTGCTTTCAG TGCAATTCTA ATTCACACGT TTAACTTTAA CATGCATGTC ATTATAATTA 240
 CTTCTTCACT AAGACACAAT ACGGCAAACC TTTCAGATTA TATTAATCTC CATAAATGAA 300
 ATAATTAACC TCATAATCAA GATTCAATGT TTCTAAATAT ATATGGACAA AATTTACACG 360
 GAAGATTAGA TACGTATATT AGTAGATTTA GTCTTTCGTT TGTGCGATAA GATTAACCAC 420
 CTCATAGATA GTAATATCAT TGTCAAATTC CTCTCGGTTT AGTCGCTAAA TTGTATCTTT 480
 TTTAAGCCTA AAAGTAGTGT ATTCGCATAT GACTTATCGT CCTAACTTTT TTTTTAATTA 540
 ACAAAAAAAT CGAAAAGAAA ATAATCTGTT AAATATTTTT TAAGTACTCC ATTAAGTTTA 600
 GTTTCTATTT AAAAAATGCT TGAAATTTGA CAGTTATGTT CAACAATTTT GAATCATGAG 660
 CGATGTCTAG ATACTCAGAA TTTAATCAAG ATGTCTTATC AAATTTGTTG TCACTCGAGG 720
 ACCCACGCAA AAGAAAAGAC TAATATGATT TTTATTTGGT CTGGATATTT TTGTAGAGGA 780
 TGAAACTAAG AGAGTGAAAG ATTCGAAATC CACAATGTTC AAGAGAGCTC AAAGCAAAAA 840
 GAAAAATGAA GATGAAGGAC TAAAGAACAA TAAGCAACTA CTTATACCCT ATTTCCATAA 900
 AGGATTCAGG TACTAGGAGA AGTTGAGGCA AGTTNNNNNN NATTGATTCA AATTTTCATT 960
 TATTTTTACA ATTTAATTCA CCTAAGTTAT TATGCATTTC TCATCATTGG TACATTTTCT 1020
 GTATAGCGTA TTTACATATA TGAAATAAAT TAAATATGTC CTCACGTTGC AAGTAGTTAA 1080
 TGAATGTCCC CACGCAAAAA AAAATCCCTC CAAATATGTC CACCTTTTCT TTTCTTTTTA 1140
 ATTCCAAAAT TACCATAAAC TTTTGGTTTA CAAAAGATTT CTAGAAATTG AGGAAGATAT 1200
 CCTAAATGAT TCATGAATCC TTCAATAATC TGAAGTTTGC GATATTTTCG ATTTTCTTCA 1260
 AGAGTTGCGA TATTTGTAAT TTGGTGACCT TAAACTTTTT TTGATAAAGA GTAAACGTTT 1320
 TTTCTTAAAA GTAAAACTTG ATTTTATGTT TTAGGGTTCT AGCTCAACTT TGTATTATAT 1380
 TTCTTGCAAA AAGAGTTCGT TAACTGCATT CTTCAACACT ATAAAGTGAT TATCAAAAAC 1440
 ATCTTCATGA ACATTAAGAA AAACAATATT TGGTTTCGGT TAGAGCTTGG TTTTGCTTGG 1500
 CTTGATTCAC ATACCCATTC TAGACTTTGG CATAAATTTG ATACGATAGA GAGTATCTAA 1560
 TGGTAATGCA GAAGGGTAAA AAAAGGAAGA GAGAAAAGGT GAGAAAGATT ACCAAAAATA 1620
 AGGAGTTTCA AAAGATGGTT CTGATGAGAA ACAGAGCCCA TCCCTCTCCT TTTCCCCTTC 1680
 CCATGAAAGA AATCGGATGG TCCTCCTTCA ATGTCCTCCA CCTACTCTTC TCTTCTTTCT 1740
 TTTTTTCTTT CTTATTATTA ACCATTTAAT TAATTTCCCC TTCAATTTCA GTTTCTAGTT 1800
 CTGTAAAAAG AAAATACACA TCTCACTTAT AGATATCCAT ATCTATTTAT ATGCATGTAT 1860
 AGAGAATAAA AAAGTGTGAG TTTCTAGGTA TGTTGAGTAT GTGCTGTTTG GACAATTGTT 1920
 AGATGATCTG TCCATTTTTT TCTTTTTTCT TCTGTGTATA AATATATTTG AGCACAAAGA 1980
 AAAACTAATA ACCTTCTGTT TTCAGCAACT AGGGTCTTAT AACCTTCAAA GAAATATTCC 2040
 TTCAATTGAA AACCCATAAA CCAAAATAGA TATTACAAAA GGAAAGAGAG ATATTTTCAA 2100
 GAACAACATA ATTAGAAAAG CAGAAGCAGC AGTTAAGTGG TACTGAGATA AATGATATAG 2160
 TTTCTCTTCA AGAACAGTTT CTCATTACCC ACCTTCTCCT TTTTGCTGAT CTATCGTAAT 2220
 CTTGAGAACT CAGGTAAGGT TGTGAATATT ATGCACCATT CATTAACCCT AAAAATAAGA 2280
 GATTTAAAAT AAATGTTTCT TCTTTCTCTG ATTCTTGTGT AACCAATTCA TGGGTTTGAT 2340
 ATGTTTCTTG GTTATTGCTT ATCAACAAAG AGATTTGATC ATTATAAAGT AGATTAATAA 2400
 CTCTTAAACA CACAAAGTTT CTTTATTTTT TAGTTACATC CCTAATTCTA GACCAGAACA 2460
 TGGATTTGAT CTATTTCTTG GTTATGTATC TTGATCAGGA AAAGGGATTT GATCATCAAG 2520
 ATTAGCCTTC TCTCTCTCTC TCTAGATATC TTTCTTGAAT TTAGAAATCT TTATTTAATT 2580
 ATTTGGTGAT GTCATATATG GATCAATGGA GGAAGGTGGG AGTAGTCACG ACGCAGAGAG 2640
 TAGCAAGAAA CTAGGGAGAG GGAAAATAGA GATAAAGAGG ATAGAGAACA CAACAAATCG 2700
 TCAAGTTACT TTCTGCAAAC GACGCAATGG TCTTCTCAAG AAAGCTTATG AACTCTCTGT 2760
 CTTGTGTGAT GCCGAAGTTG CCCTCGTCAT CTTCTCCACT CGTGGCCGTC TCTATGAGTA 2820
 CGCCAACAAC AGGTACGCTT CTCCTACTCT ATTTCTTGAT CTTGTTTTCT TAATTTTAAC 2880
 TAAACAAGAT CCTAGTTCAA ATGATAACAA AGTGGGGATT GAGAGCCAAG ATTAGGGTTT 2940
 GGTTAATTTA GAAAACCAGA TTTCACTTGT TGATACATTT AATATCTCTC TAGCTAGATT 3000
 TAGTACTCTC TCCTCTATAT ATGTGTGGGT GTGTGTGTAA GTGTGTATAT GTATGCAAAT 3060
 GCAAGAAGAA GAAGAAAAAG TTATCTTGTC TTCTCAAATT CTGATCAGCT TTGACCTTAG 3120
 TTTCACTCTT TTTTCTGCAA ATCATTTGAA CCTGATGCAT GTCAGTTTCT ACAATACACT 3180
 TTTAATTTTG ACGGCCCATC AAATTTCCTA GGGTTTACTT CAGTGAACAA AATTGGGTTC 3240
 TTGACACGAT TTAGCATGTA TATATAAAAA TAGGGGATGA TCAAGACTTA TGTAACCTCT 3300
 GTCTGGTGAA ACTAGGGACA AAGTCTACTG ATGAGTTGTC ACTAGGGATC CATTTGATCA 3360
 TTTAATCCCA ACAAAAATGA AACAAAATTT TGAGAATTTA TATGCTGAAG TTTTTCAACC 3420
 CTCTTTTTTA AATAACTTTA TATTATGTAG ATTTGTATTT AGGGTAATTT GTCCAACTAG 3480
 AAGTCCTAAA AATCAATAAA CACACGGATG ACTTTGTCTA ACATTGTATC AGTCATCAAA 3540
 TGTAAAATTG TACAAATAAT GAAATTAAAG ATTTAGTCTC TTTTATTTTT TTTGTTTAGG 3600
 GTGTATATAT ATATATATAT GTATATTTGT TGCATTGATA TATCAATGAG AGGGAGAGAA 3660
 CTCAGAGAAG TGTCGGAAAT TAAAATGGTA CGAGCCAATT GGAATCTCTG GCATTCTGAG 3720
 CTTCATTTGT TTGTTATTAG AAAAAAAAAA AAAAAATCCT TTAAAGATAC CTTCATGATG 3780
 ACATTGAATC ATGTAATATA CACGATACAT GGTCTAATTC CTCCTCAAAC CCTAATTACC 3840
 AATTTCGAAA CCATAATATT TACTAGTATG TTTATATATC CTTACTTTAA GACATTGTTT 3900
 GTTTATAATA CCTTGTGAAT TAAGAAAAAA AAAAAAAAAC TTGTGGATCT ATTCAAGCCA 3960
 TGTGTTAGAA TAAATTTATA AATTTTCTCC TCGTACTGGT CAGATATTGG TCCAAACTCC 4020
 AAAGCCTTCC CTTTTCAGGA AAAAAAACAT TTCGAAATTA ACTCTAATTA ATCAAGAATT 4080
 TCCTACAATG TATACATCTA ATGTTTTTTC CGCGATCTTA CTTATTAGTG TGAGGGGTAC 4140
 AATTGAAAGG TACAAGAAAG CTTGTTCCGA TGCCGTCAAC CCTCCTTCCG TCACCGAAGC 4200
 TAATACTCAG GTACCAATTT ATATTGTTTG ATTCTCTTTG TTTTATCTTC TTCTTTTCAT 4260
 TATATATATG ATCAACAAAA AATATAACCT ACAAAAAGAG AGAGTTCAAG GAAATGCATT 4320
 GAAACGGTTT CGTTATGGTG TTTGAATACA TGGATTTTTG AAGTACTATC AGCAAGAAGC 4380
 CTCTAAGCTT CGGAGGCAGA TTCGAGATAT TCAGAATTCA AATAGGTAAT TCATTAACTT 4440
 TTCATGAACT CTTCGATTTG GTATTAGGTC ACTTAATTTG GTGTCGGTCC AAAAGTCCGC 4500
 TTGTAGTTTT CTTTAGAAGT TGTTTTGTTT AATGTTCATG TTTACAAATT GAAGGCATAT 4560
 TGTTGGGGAA TCACTTGGTT CCTTGAACTT CAAGGAACTC AAAAACCTAG AAGGACGTCT 4620
 TGAAAAAGGA ATCAGCCGTG TCCGCTCCAA AAAGGTAAAA TCTACGTTGC TCTCTCTCTG 4680
 TGTCTCTGTC TCTCTCTCTA TATATAGTCC CTTAGTTTAT ATAGTTCATC ACCCTTTTGT 4740
 GAGAATTTTG CAGAATGAGC TGTTAGTGGC AGAGATAGAG TATATGCAGA AGAGGGTAAG 4800
 AACGTTTCTC CCATTCCAAG TAATTAGATC TTTCTTCGTC TTTGTGAGGG TTTGAGTTTT 4860
 CCCATAAATC ATGTGTAGGA AATGGAGTTG CAACACAATA ACATGTACCT GCGAGCAAAG 4920
 GTTAGCCACG TTCTGTTCCA AATCTTAATC TCAATATCTA CTCTTTTCTT CATTGTATAA 4980
 CTAAGATAAC GTGAATAACA AGAAAACTTT TGTTTTTGGG TTTAATAGAT AGCCGAAGGC 5040
 GCCAGATTGA ATCCGGACCA GCAGGAATCG AGTGTGATAC AAGGGACGAC AGTTTACGAA 5100
 TCCGGTGTAT CTTCTCATGA CCAGTCGCAG CATTATAATC GGAACTATAT TCCGGTGAAC 5160
 CTTCTTGAAC CGAATCAGCA ATTCTCCGGC CAAGACCAAC CTCCTCTTCA ACTTGTGTAA 5220
 CTCAAAACAT GATAACTTGT TTCTTCCCCT CATAACGATT AAGAGAGAGA CGAGAGAGTT 5280
 CATTTTATAT TTATAACGCG ACTGTGTATT CATAGTTTAG GTTCTAATAA TGATAATAAC 5340
 AAAACTGTTG TTTCTTTGCT TAATTACATC AACATTTAAA TCCAAAGTTC TAAAACACGT 5400
 CGAGATCCAA AGTTTGTCAT ACAAGATTAG ACGCATACAC GATCAGTTAA TAGATTTTAA 5460
 GTGCCTTTTA ATATTTACAT ATAGTTGCAG CTTCGATTAG ATCATGTCCA CCAAACACTC 5520
 ACAATTAGAG ACAAGCAAAA CTATAAACAT TGATCATAAA ATGATTACAA CATGTCCATA 5580
 AATTAATTAT GGATTACAAA AATAAAAACT TACAAAAGAT CT 5622
 (2) INFORMATION FOR SEQ ID NO: 4:
 (i) SEQUENCE CHARACTERISTICS:
 (A) LENGTH: 6138 base pairs
 (B) TYPE: nucleic acid
 (C) STRANDEDNESS: unknown
 (D) TOPOLOGY: unknown
 (vi) ORIGINAL SOURCE:
 (A) ORGANISM: not provided
 (ix) FEATURE:
 (A) NAME/KEY: misc_feature
 (B) LOCATION: 1..6138
 (D) OTHER INFORMATION: /label= AGL5_promoter
 /note= "Nucleotide sequence of the AGL5 promoter."
 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:4
 GAATTCGTAA CAGAATTTAG TGAATAATAT TGTAATTACC AGGCAAGGAC TCTCCAAACG 60
 GATAGCTCGA ATATCGTTAT TAAAGAGTAA ATGATCCAAT ATGTAAGCCA TTGTTGATCA 120
 TCTAACATTG TTGGACTCTC TATTGCTCGA AATGATGCAT ACCTAATCAT TTATTCAGTT 180
 AACTATCAAG TTGCATTTGT AAAAACCAAA CATTTAAATT CAGATTTGAT ATCACTTACA 240
 GAGGATAGAG AAGCATGACT CCAGGCCTGC ATGCAACAAG AAAAAGGAAG AAAATAATGT 300
 TAAAAATTTG ACAAATATAG TGTTTATTTT TATTATATGA GACAGAATTT GAATAAAATC 360
 CTACCCAACT AGAGCATCAA AACGTTTTGC AATCGCAATA ATGAAACCCA TTTTCTTTTT 420
 GAGTTTTTAC TCTTCTTTCA ACAGAAACTT TCTCAAACGT CTTTAGCACT GTGACGTTAG 480
 ATATATACAC AAAAGCTTGA AATTTCTTCA AGCAAAAGAA TCTTTGTGGG AGTTAAGGCA 540
 ACAAGCCAGG TAAAGAATCT CCAACGCATT GTTACGTTTT CATGAACCTA TTTATTATAT 600
 GTTCTAAGAA AGAAAAAAAT ATCTCAAAGT AAACGTTGGA AATTTTCTGA TGAAGGGAAA 660
 TCCAAAGTCT TGGGTTTAGT ATCCCTATGA ATGGTATTTG GAATATGTTT TCGTCAAAAC 720
 AAAAGATTCT TTTCTTTTTC ACAAGAGTTA GTGATCAATA ACTTATGCAC TAATTAATGA 780
 GATTGGACGT ATACACAATT TGATTATGAT ACTTGAGTAA AAATCACCTG TCCTTTAATT 840
 TGGAAATCTC TCTTTCTTAC CCATTTATAT ACTACTTCTT TTCATTAAAA TTAAATTTCA 900
 ATTATCAATC ATCGTTCAAT TTGATAAAGA TTTAACATTT TTTGTCACAG GGCTAGTAAA 960
 AGCAATCTTT ACATAATTCA TCTTTCTTAC ATATATATAT TACCTTTTTC TTCATTAGTA 1020
 TTCTATTTGA TTATGATTAT TTTGTCATAA AGCTAGTAAA TTAAACACTC GATATGAGAA 1080
 TTATATTACT TCACGCTAAT TAACTCTTAA CACAACAAGA ACTAGTGCAT ATTCAACTTT 1140
 CAAAGCATAT ACTATATATT GAGAATATAG ACCACGAAAG TCAATCAAAA GACCTACCAG 1200
 CTCTCATCAA GTTCTTTCTT GAAATGATTT TGCAGAATTT CCAACTTAAT TAATTCGACA 1260
 TGAATGTGAA AATGTGTGTT GCTCGTTAAG AAAATTGAAT AGAAGTACAA TGAAAATGAT 1320
 GAGGAATGGG CAAAACACAA AAGAGTTTCC TTTCGTAACT ACAATTAATT AATGCAAATC 1380
 TGAGAAAGGG TTCATGGATA ATGACTACAC ACATGATTAG TCATTCCCCG TGGGCTCTCT 1440
 GCTTTCATTT ACTTTATTAG TTTCATCTTC TCTAATTATA TTGTCGCATA TATGATGCAG 1500
 TTCTTTTGTC TAAATTACGT AATATGATGT AATTAATTAT CAAAATAAAT ATTCAAATTG 1560
 CCGTTGGACT AACCTAATGT CCAAGATTAA GACTTGAACA TAAGAATTTT GGAAAAACTA 1620
 AACCAGTTAT AATATATACT CTTAAATTGC CATTTCTGAA CACAACCAAA TAATAATATA 1680
 TACTATTTAC AGTTTTTTTT AATTGGCAAG AACACTGAAA TCTTATTCAT TGTCTCGCTT 1740
 GGTAGTTGAC AAGTTATAAC ACTCATATTC ATATAACCCC ATTCTAACGT TGACGACGAA 1800
 CACTCATATA AACCACCCAA ATTCTTAGCA TATTAGCTAA ATATTGGTTT AATTGGAAAT 1860
 ATTTTTTTTA TATATAAAAT GCCAGGTAAA TATTAACGAC ATGCAATGTA TATAGGAGTA 1920
 GGGCAATAAA AAGAAAAGGA GAATAAAAAG GGATTACCAA AAAAGGAAAG TTTCCAAAAG 1980
 GTGATTCTGA TGAGAAACAG AGCCCATACC TCTCTTTTTT CCTCTAAACA TGAAAGAAAA 2040
 ATTGGATGGT CCTCCTTCAA TGCTCTCTCC CCACCCAATC CAAACCCAAC TGTCTTCTTT 2100
 CTTTCTTTTT TCTTCTTTCT AATTTGATAT TTTCTACCAC TTAATTCCAA TCAATTTCAA 2160
 ATTTCAATCT AAATGTATGC ATATAGAATT TAATTAAAAG AATTAGGTGT GTGATATTTG 2220
 AGAAAATGTT AGAAGTAATG GTCCATGTTC TTTCTTTCTT TTTCCTTCTA TAACACTTCA 2280
 GTTTGAAAAA AAACTACCAA ACCTTCTGTT TTCTGCAAAT GGGTTTTTAA ATACTTCCAA 2340
 AGAAATATTC CTCTAAAAGA AATTATAAAC CAAAACAGAA ACCAAAAACA AAAAATAAAG 2400
 TTGAAGCAGC AGTTAAGTGG TACTGAGATA ATAAGAATAG TATCTTTAGG CCAATGAACA 2460
 AATTAACTCT CTCATAATTC ATCTTCCCAT CCTCACTTCT CTTTCTTTCT GATATAATTA 2520
 ATCTTGCTAA GCCAGGTATG GTTATTGATG ATTTACACTT TTTTTTAAAA GTTTCTTCCT 2580
 TTTCTCCAAT CAAATTCTTC AGTTAATCCT TATAAACCAT TTCTTTAATC CAAGGTGTTT 2640
 GAGTGCAAAA GGATTTGATC TATTTCTCTT GTGTTTATAC TTCAGCTAGG GCTTATAGAA 2700
 ATGGAGGGTG GTGCGAGTAA TGAAGTAGCA GAGAGCAGCA AGAAGATAGG GAGAGGGAAG 2760
 ATAGAGATAA AGAGGATAGA GAACACTACG AATCGTCAAG TCACTTTCTG CAAACGACGC 2820
 AATGGTTTAC TCAAGAAAGC TTATGAGCTC TCTGTCTTGT GTGACGCTGA GGTTGCTCTT 2880
 GTCATCTTCT CCACTCGAGG CCGTCTCTAC GAGTACGCCA ACAACAGGTA CACATCTTTT 2940
 AGCTAGATCT TGATTTTGTT GAATTTTTTT TCTAGAATAA AGTTTCGACT CTTCTGGTGG 3000
 GTTTTTCAAT CTTTATGGTC TCTTTATAGT TTTTTTCCTT AGTTTCTCTG AAGCTCAAAT 3060
 CTCTTTAAAA ATCCCCAAAA TTAGGGTTTG TTTAAAACTA GGGAACCCTA CTTTAACTTC 3120
 TTTCTCTTAG TAAAAAAGCA GTGAGGGTCT TCTCTGATCA TTAATTAGCA TCCCCCATAC 3180
 CTTGTTCCAG TCACTTTTTC TCCACAAATC CTTATAACAG TATCTATATA TGTATCTATT 3240
 TATGTCAGTT TGTACAAGAC ACTTCGATCA ATTTGATGAC CCATCAAGTT TTATTTCTGC 3300
 AGATTGATCA TTAGGTTTCC ATCATAGTAA TGAAAAAGTA GGGTTCTTGA TAAAATTATA 3360
 ATAATATATA TTATTTGGCT ATATAAAAAA GCTATGTAGA TTCCTTAAAA ATTGATTCAC 3420
 TAGGGAGAGA CTAGTAGGTG TTTGTCTTCT GACACTTCTC TAATCTTTTG GTGAATCCTT 3480
 TTGTTAAATC AAGAAAATGA ATCAGGGACA AAGCTTATTG TTGAGTCACT TAATTAATCA 3540
 TCCGATCCAT CAATCAAGAA AAATAACGAA ACAGAAAATT TTGATTTTTG ATTGTTATTT 3600
 TCTCCACTTC AAGTTGGGGA CTTGTCATTT CCGTTTTTCT ATACGTTTCC AGCTATTAAC 3660
 AGCTCATGTT CATTTCACCA TTTTGATTAT TTGTCTGCTT TTTAAAGATA AATGTTTTCA 3720
 AAAATATTGT TTTTATTTGC TTGGCTAGTT AATACTATAA TTGAGGTTGA TGTATGACTA 3780
 TAATCTATAA GTCAAGTCTC ATATCATGGA TCTAAGTTAA AACTAGTAAA TTTGTAGTTT 3840
 CAATGTGAAC TTTCACAACG ACTAAAGAAC TGATCTGAAG TTTATAATGG ACATGACTAA 3900
 TTTGATTAAC AAAAGAGGAA TGCATTATGT ATGTAGAAAC ATGTGATATA TATATGTTTC 3960
 TATTATCAAA AGTGTAGTTA ACTTTCTTAT TTCAAACACC CTCATGCTTT AGTAGTATCT 4020
 TACTTTTGAC ATTTCTCAAC TTCAGCTTTC CATTATACAA CAGCACAATG TAAATTACTT 4080
 GTATATGAAT ATGAAAGCAT AACGTTATGC AAAGATTTCT AGCTTTTCTT TTTCTGTTTT 4140
 GCAAAAGATT TACAAATATC ATGTTCTTGG TAAAAACATA CTTGCCTCAG CCACATATGC 4200
 ATGTAAATGT AATGTTCAAA TATTAATTCA GGAAAAACAA AGAAGAAGCA AAATTAGCTT 4260
 CTAGAGTAGG GAATCTATTG ACTTGACCTG AAAATCACTT CTTTTTCTTA AAGCCTAGTA 4320
 GTGAATTTTT TAATCTAATT AGGCCAAAAT ATATACTAGC CTAAAATATA ATTTGGATTT 4380
 TGTGTCGTAC ATAAATTGGG ACCAATTCCA ATTAACTAAG AGCATATGCA ATTCAAATTC 4440
 TTTTTATTTT CTTCTCCGAT TTGCTACTTC TTTCTTTTGT ATGTTTTCAA ATTAGGATTA 4500
 CACTTTTTTG GGGAAGTACA CATTAGGGTC TTCTCGAACT TTGATTATAC ATATATATAT 4560
 ATATATATAT ATATAACTTT GTGAGATGTC ACTGTTAATA GATAATAGGC AATAACAATA 4620
 ATATCCAAAA AAGAAGGCGC AAACAAATCA TATACTATAT GGTACTGGTC CATTCACTAT 4680
 TTTGTCGGTT GAATTTAAGG TTTGGCGTAC AAACTTTGTT TCAAACCTTT ATTATTCCGT 4740
 CTTTCTGTGT GTTTTGTATA TCCAGAAGAT AAAAATATCA ATTTCTTTAA CGACTTCATA 4800
 TATATATATA TATATATATA TATATATATT TTTCTCTTCT GGTTTTAGTG TTTGAATCCA 4860
 ACAGTTATAG TTTCGTGTGT CTTTGTTTTA CTTGTGGTGG TTTAAGTTTG AGATTTTCAC 4920
 CGATTGCATC TATTTACATA TATAGCTACC ACAAAAAAGA TTGCATTTTA AAATCTTTTC 4980
 CTTTGTGTGA ATGTTGATGA AGTGTGAGAG GAACAATAGA AAGGTACAAG AAAGCTTGCT 5040
 CCGACGCCGT TAACCCTCCG ACCATCACCG AAGCTAATAC TCAGGTTAGC TTTTAATTAA 5100
 TACACCTAGC TAGCTAGTTC GTTAATTACT TAATTTCTTC TTCTTTTAGT TATCTGACCT 5160
 TTTTTTCACC TCTTGTAACA ATGATGGGAT CGAAATTGAT GAAGTACTAT CAGCAAGAGG 5220
 CGTCTAAACT CCGGAGACAG ATTCGGGACA TTCAGAATTT GAACAGACAC ATTCTTGGTG 5280
 AATCTCTTGG TTCCTTGAAC TTTAAGGAAC TCAAGAACCT TGAAAGTAGG CTTGAGAAAG 5340
 GAATCAGTCG TGTCCGATCC AAGAAGGTAC ATCACTAACT CTCCATCAAT CTCCTTATCA 5400
 TTGAATATAT ATCCATCTGA TTCTTGCCCG TTATATTTGG TTTTTCTCTC CAGCACGAGA 5460
 TGTTAGTTGC AGAGATTGAA TACATGCAAA AAAGGGTAAA AGTAAAACCT ATCTTCCTTC 5520
 ACAATGAACT ACCCCTACTT TATTAGCAAC TTCTCTTTCT GATGATCATC TTTTTTATTT 5580
 TCTGTTGTCG CTTGCATTGT AGGAAATCGA GCTGCAAAAC GATAACATGT ATCTCCGCTC 5640
 CAAGGTTTTA TACATAACTC TTTTTGGCAT TTTTGATCAT CATTTTTTTC CGGTAGACAA 5700
 TCTCTTGATG TGCAAATTCT AAATATCTCT GCAGATTACT GAAAGAACAG GTCTACAGCA 5760
 ACAAGAATCG AGTGTGATAC ATCAAGGGAC AGTTTACGAG TCGGGTGTTA CTTCTTCTCA 5820
 CCAGTCGGGG CAGTATAACC GGAATTATAT TGCGGTTAAC CTTCTTGAAC CGAATCAGAA 5880
 TTCCTCCAAC CAAGACCAAC CACCTCTGCA ACTTGTTTGA TTCAGTCTAA CATAAGCTTC 5940
 TTTCCTCAGC CTGAGATCGA TCTATAGTGT CACCTAAATG CGGCCGCGTC CCTCAACATC 6000
 TAGTCGCAAG CTGAGGGGAA CCACTAGTGT CATACGAACC TCCAAGAGAC GGTTACACAA 6060
 ACGGGTACAT TGTTGATGTC ATGTATGACA ATCGCCCAAG TAAGTATCCA GCTGTGTTCA 6120
 GAACGTACGT CCGAATTC 6138
 (2) INFORMATION FOR SEQ ID NO: 5:
 (i) SEQUENCE CHARACTERISTICS:
 (A) LENGTH: 896 base pairs
 (B) TYPE: nucleic acid
 (C) STRANDEDNESS: single
 (D) TOPOLOGY: linear
 (ii) MOLECULE TYPE: cDNA
 (vi) ORIGINAL SOURCE:
 (A) ORGANISM: not provided
 (ix) FEATURE:
 (A) NAME/KEY: CDS
 (B) LOCATION: 7..753
 (ix) FEATURE:
 (A) NAME/KEY: misc_feature
 (B) LOCATION: 896
 (D) OTHER INFORMATION: /note= "There is a poly(A) tail at
 the end of the cDNA sequence."
 (ix) FEATURE:
 (A) NAME/KEY: misc_feature
 (B) LOCATION: 1..896
 (D) OTHER INFORMATION: /note= "AGL1 cDNA and deduced
 protein sequences."
 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:5
 GGATCA ATG GAG GAA GGT GGG AGT AGT CAC GAC GCA GAG AGT AGC AAG 48
 Met Glu Glu Gly Gly Ser Ser His Asp Ala Glu Ser Ser Lys
 1 5 10
 AAA CTA GGG AGA GGG AAA ATA GAG ATA AAG AGG ATA GAG AAC ACA ACA 96
 Lys Leu Gly Arg Gly Lys Ile Glu Ile Lys Arg Ile Glu Asn Thr Thr
 15 20 25 30
 AAT CGT CAA GTT ACT TTC TGC AAA CGA CGC AAT GGT CTT CTC AAG AAA 144
 Asn Arg Gln Val Thr Phe Cys Lys Arg Arg Asn Gly Leu Leu Lys Lys
 35 40 45
 GCT TAT GAA CTC TCT GTC TTG TGT GAT GCC GAA GTT GCC CTC GTC ATC 192
 Ala Tyr Glu Leu Ser Val Leu Cys Asp Ala Glu Val Ala Leu Val Ile
 50 55 60
 TTC TCC ACT CGT GGC CGT CTC TAT GAG TAC GCC AAC AAC AGT GTG AGG 240
 Phe Ser Thr Arg Gly Arg Leu Tyr Glu Tyr Ala Asn Asn Ser Val Arg
 65 70 75
 GGT ACA ATT GAA AGG TAC AAG AAA GCT TGT TCC GAT GCC GTC AAC CCT 288
 Gly Thr Ile Glu Arg Tyr Lys Lys Ala Cys Ser Asp Ala Val Asn Pro
 80 85 90
 CCT TCC GTC ACC GAA GCT AAT ACT CAG TAC TAT CAG CAA GAA GCC TCT 336
 Pro Ser Val Thr Glu Ala Asn Thr Gln Tyr Tyr Gln Gln Glu Ala Ser
 95 100 105 110
 AAG CTT CGG AGG CAG ATT CGA GAT ATT CAG AAT TCA AAT AGG CAT ATT 384
 Lys Leu Arg Arg Gln Ile Arg Asp Ile Gln Asn Ser Asn Arg His Ile
 115 120 125
 GTT GGG GAA TCA CTT GGT TCC TTG AAC TTC AAG GAA CTC AAA AAC CTA 432
 Val Gly Glu Ser Leu Gly Ser Leu Asn Phe Lys Glu Leu Lys Asn Leu
 130 135 140
 GAA GGA CGT CTT GAA AAA GGA ATC AGC CGT GTC CGC TCC AAA AAG AAT 480
 Glu Gly Arg Leu Glu Lys Gly Ile Ser Arg Val Arg Ser Lys Lys Asn
 145 150 155
 GAG CTG TTA GTG GCA GAG ATA GAG TAT ATG CAG AAG AGG GAA ATG GAG 528
 Glu Leu Leu Val Ala Glu Ile Glu Tyr Met Gln Lys Arg Glu Met Glu
 160 165 170
 TTG CAA CAC AAT AAC ATG TAC CTG CGA GCA AAG ATA GCC GAA GGC GCC 576
 Leu Gln His Asn Asn Met Tyr Leu Arg Ala Lys Ile Ala Glu Gly Ala
 175 180 185 190
 AGA TTG AAT CCG GAC CAG CAG GAA TCG AGT GTG ATA CAA GGG ACG ACA 624
 Arg Leu Asn Pro Asp Gln Gln Glu Ser Ser Val Ile Gln Gly Thr Thr
 195 200 205
 GTT TAC GAA TCC GGT GTA TCT TCT CAT GAC CAG TCG CAG CAT TAT AAT 672
 Val Tyr Glu Ser Gly Val Ser Ser His Asp Gln Ser Gln His Tyr Asn
 210 215 220
 CGG AAC TAT ATT CCG GTG AAC CTT CTT GAA CCG AAT CAG CAA TTC TCC 720
 Arg Asn Tyr Ile Pro Val Asn Leu Leu Glu Pro Asn Gln Gln Phe Ser
 225 230 235
 GGC CAA GAC CAA CCT CCT CTT CAA CTT GTG TAACTCAAAA CATGATAACT 770
 Gly Gln Asp Gln Pro Pro Leu Gln Leu Val
 240 245
 TGTTTCTTCC CCTCATAACG ATTAAGAGAG AGACGAGAGA GTTCATTTTA TATTTATAAC 830
 GCGACTGTGT ATTCATAGTT TAGGTTCTAA TAATGATAAT AACAAAACTG TTGTTTCTTT 890
 GCTTCA 896
 (2) INFORMATION FOR SEQ ID NO: 6:
 (i) SEQUENCE CHARACTERISTICS:
 (A) LENGTH: 248 amino acids
 (B) TYPE: amino acid
 (D) TOPOLOGY: linear
 (ii) MOLECULE TYPE: protein
 (vi) ORIGINAL SOURCE:
 (A) ORGANISM: not provided
 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:6
 Met Glu Glu Gly Gly Ser Ser His Asp Ala Glu Ser Ser Lys Lys Leu
 1 5 10 15
 Gly Arg Gly Lys Ile Glu Ile Lys Arg Ile Glu Asn Thr Thr Asn Arg
 20 25 30
 Gln Val Thr Phe Cys Lys Arg Arg Asn Gly Leu Leu Lys Lys Ala Tyr
 35 40 45
 Glu Leu Ser Val Leu Cys Asp Ala Glu Val Ala Leu Val Ile Phe Ser
 50 55 60
 Thr Arg Gly Arg Leu Tyr Glu Tyr Ala Asn Asn Ser Val Arg Gly Thr
 65 70 75 80
 Ile Glu Arg Tyr Lys Lys Ala Cys Ser Asp Ala Val Asn Pro Pro Ser
 85 90 95
 Val Thr Glu Ala Asn Thr Gln Tyr Tyr Gln Gln Glu Ala Ser Lys Leu
 100 105 110
 Arg Arg Gln Ile Arg Asp Ile Gln Asn Ser Asn Arg His Ile Val Gly
 115 120 125
 Glu Ser Leu Gly Ser Leu Asn Phe Lys Glu Leu Lys Asn Leu Glu Gly
 130 135 140
 Arg Leu Glu Lys Gly Ile Ser Arg Val Arg Ser Lys Lys Asn Glu Leu
 145 150 155 160
 Leu Val Ala Glu Ile Glu Tyr Met Gln Lys Arg Glu Met Glu Leu Gln
 165 170 175
 His Asn Asn Met Tyr Leu Arg Ala Lys Ile Ala Glu Gly Ala Arg Leu
 180 185 190
 Asn Pro Asp Gln Gln Glu Ser Ser Val Ile Gln Gly Thr Thr Val Tyr
 195 200 205
 Glu Ser Gly Val Ser Ser His Asp Gln Ser Gln His Tyr Asn Arg Asn
 210 215 220
 Tyr Ile Pro Val Asn Leu Leu Glu Pro Asn Gln Gln Phe Ser Gly Gln
 225 230 235 240
 Asp Gln Pro Pro Leu Gln Leu Val
 245
 (2) INFORMATION FOR SEQ ID NO: 7:
 (i) SEQUENCE CHARACTERISTICS:
 (A) LENGTH: 959 base pairs
 (B) TYPE: nucleic acid
 (C) STRANDEDNESS: single
 (D) TOPOLOGY: linear
 (ii) MOLECULE TYPE: cDNA
 (vi) ORIGINAL SOURCE:
 (A) ORGANISM: not provided
 (ix) FEATURE:
 (A) NAME/KEY: CDS
 (B) LOCATION: 78..818
 (ix) FEATURE:
 (A) NAME/KEY: misc_feature
 (B) LOCATION: 1..959
 (D) OTHER INFORMATION: /note= "AGL5 cDNA and deduced
 protein sequences."
 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:7
 GAATTCATCT TCCCATCCTC ACTTCTCTTT CTTTCTGATC ATAATTAATC TTGCTAAGCC 60
 AGCTAGGGCT TATAGAA ATG GAG GGT GGT GCG AGT AAT GAA GTA GCA GAG 110
 Met Glu Gly Gly Ala Ser Asn Glu Val Ala Glu
 1 5 10
 AGC AGC AAG AAG ATA GGG AGA GGG AAG ATA GAG ATA AAG AGG ATA GAG 158
 Ser Ser Lys Lys Ile Gly Arg Gly Lys Ile Glu Ile Lys Arg Ile Glu
 15 20 25
 AAC ACT ACG AAT CGT CAA GTC ACT TTC TGC AAA CGA CGC AAT GGT TTA 206
 Asn Thr Thr Asn Arg Gln Val Thr Phe Cys Lys Arg Arg Asn Gly Leu
 30 35 40
 CTC AAG AAA GCT TAT GAG CTC TCT GTC TTG TGT GAC GCT GAG GTT GCT 254
 Leu Lys Lys Ala Tyr Glu Leu Ser Val Leu Cys Asp Ala Glu Val Ala
 45 50 55
 CTT GTC ATC TTC TCC ACT CGA GGC CGT CTC TAC GAG TAC GCC AAC AAC 302
 Leu Val Ile Phe Ser Thr Arg Gly Arg Leu Tyr Glu Tyr Ala Asn Asn
 60 65 70 75
 AGT GTG AGA GGA ACA ATA GAA AGG TAC AAG AAA GCT TGC TCC GAC GCC 350
 Ser Val Arg Gly Thr Ile Glu Arg Tyr Lys Lys Ala Cys Ser Asp Ala
 80 85 90
 GTT AAC CCT CCG ACC ATC ACC GAA GCT AAT ACT CAG TAC TAT CAG CAA 398
 Val Asn Pro Pro Thr Ile Thr Glu Ala Asn Thr Gln Tyr Tyr Gln Gln
 95 100 105
 GAG GCG TCT AAA CTC CGG AGA CAG ATT CGG GAC ATT CAG AAT TTG AAC 446
 Glu Ala Ser Lys Leu Arg Arg Gln Ile Arg Asp Ile Gln Asn Leu Asn
 110 115 120
 AGA CAC ATT CTT GGT GAA TCT CTT GGT TCC TTG AAC TTT AAG GAA CTC 494
 Arg His Ile Leu Gly Glu Ser Leu Gly Ser Leu Asn Phe Lys Glu Leu
 125 130 135
 AAG AAC CTT GAA AGT AGG CTT GAG AAA GGA ATC AGT CGT GTC CGA TCC 542
 Lys Asn Leu Glu Ser Arg Leu Glu Lys Gly Ile Ser Arg Val Arg Ser
 140 145 150 155
 AAG AAG CAC GAG ATG TTA GTT GCA GAG ATT GAA TAC ATG CAA AAA AGG 590
 Lys Lys His Glu Met Leu Val Ala Glu Ile Glu Tyr Met Gln Lys Arg
 160 165 170
 GAA ATC GAG CTG CAA AAC GAT AAC ATG TAT CTC CGC TCC AAG ATT ACT 638
 Glu Ile Glu Leu Gln Asn Asp Asn Met Tyr Leu Arg Ser Lys Ile Thr
 175 180 185
 GAA AGA ACA GGT CTA CAG CAA CAA GAA TCG AGT GTG ATA CAT CAA GGG 686
 Glu Arg Thr Gly Leu Gln Gln Gln Glu Ser Ser Val Ile His Gln Gly
 190 195 200
 ACA GTT TAC GAG TCG GGT GTT ACT TCT TCT CAC CAG TCG GGG CAG TAT 734
 Thr Val Tyr Glu Ser Gly Val Thr Ser Ser His Gln Ser Gly Gln Tyr
 205 210 215
 AAC CGG AAT TAT ATT GCG GTT AAC CTT CTT GAA CCG AAT CAG AAT TCC 782
 Asn Arg Asn Tyr Ile Ala Val Asn Leu Leu Glu Pro Asn Gln Asn Ser
 220 225 230 235
 TCC AAC CAA GAC CAA CCA CCT CTG CAA CTT GTT TGATTCAGTC TAACATAAGC 835
 Ser Asn Gln Asp Gln Pro Pro Leu Gln Leu Val
 240 245
 TTCTTTCCTC AGCCTGAGAT CGATCTATAG TGTCACCTAA ATGCGGCCGC GTCCCTCAAC 895
 ATCTAGTCGC AAGCTGAGGG GAACCACTAG TGTCATACGA ACCTCCAAGA GACGGTTACA 955
 CAAA 959
 (2) INFORMATION FOR SEQ ID NO: 8:
 (i) SEQUENCE CHARACTERISTICS:
 (A) LENGTH: 246 amino acids
 (B) TYPE: amino acid
 (D) TOPOLOGY: linear
 (ii) MOLECULE TYPE: protein
 (vi) ORIGINAL SOURCE:
 (A) ORGANISM: not provided
 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:8
 Met Glu Gly Gly Ala Ser Asn Glu Val Ala Glu Ser Ser Lys Lys Ile
 1 5 10 15
 Gly Arg Gly Lys Ile Glu Ile Lys Arg Ile Glu Asn Thr Thr Asn Arg
 20 25 30
 Gln Val Thr Phe Cys Lys Arg Arg Asn Gly Leu Leu Lys Lys Ala Tyr
 35 40 45
 Glu Leu Ser Val Leu Cys Asp Ala Glu Val Ala Leu Val Ile Phe Ser
 50 55 60
 Thr Arg Gly Arg Leu Tyr Glu Tyr Ala Asn Asn Ser Val Arg Gly Thr
 65 70 75 80
 Ile Glu Arg Tyr Lys Lys Ala Cys Ser Asp Ala Val Asn Pro Pro Thr
 85 90 95
 Ile Thr Glu Ala Asn Thr Gln Tyr Tyr Gln Gln Glu Ala Ser Lys Leu
 100 105 110
 Arg Arg Gln Ile Arg Asp Ile Gln Asn Leu Asn Arg His Ile Leu Gly
 115 120 125
 Glu Ser Leu Gly Ser Leu Asn Phe Lys Glu Leu Lys Asn Leu Glu Ser
 130 135 140
 Arg Leu Glu Lys Gly Ile Ser Arg Val Arg Ser Lys Lys His Glu Met
 145 150 155 160
 Leu Val Ala Glu Ile Glu Tyr Met Gln Lys Arg Glu Ile Glu Leu Gln
 165 170 175
 Asn Asp Asn Met Tyr Leu Arg Ser Lys Ile Thr Glu Arg Thr Gly Leu
 180 185 190
 Gln Gln Gln Glu Ser Ser Val Ile His Gln Gly Thr Val Tyr Glu Ser
 195 200 205
 Gly Val Thr Ser Ser His Gln Ser Gly Gln Tyr Asn Arg Asn Tyr Ile
 210 215 220
 Ala Val Asn Leu Leu Glu Pro Asn Gln Asn Ser Ser Asn Gln Asp Gln
 225 230 235 240
 Pro Pro Leu Gln Leu Val
 245
 (2) INFORMATION FOR SEQ ID NO: 9:
 (i) SEQUENCE CHARACTERISTICS:
 (A) LENGTH: 27 base pairs
 (B) TYPE: nucleic acid
 (C) STRANDEDNESS: single
 (D) TOPOLOGY: linear
 (vi) ORIGINAL SOURCE:
 (A) ORGANISM: not provided
 (ix) FEATURE:
 (A) NAME/KEY: misc_feature
 (B) LOCATION: 1..27
 (D) OTHER INFORMATION: /note= "Primer AGL8 5-4"
 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:9
 CCGTCGACGA TGGGAAGAGG TAGGGTT 27
 (2) INFORMATION FOR SEQ ID NO: 10:
 (i) SEQUENCE CHARACTERISTICS:
 (A) LENGTH: 20 base pairs
 (B) TYPE: nucleic acid
 (C) STRANDEDNESS: single
 (D) TOPOLOGY: linear
 (vi) ORIGINAL SOURCE:
 (A) ORGANISM: not provided
 (ix) FEATURE:
 (A) NAME/KEY: misc_feature
 (B) LOCATION: 1..20
 (D) OTHER INFORMATION: /note= "Primer OAM14."
 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:10
 AATCATTACC AAGATATGAA 20
 (2) INFORMATION FOR SEQ ID NO: 11:
 (i) SEQUENCE CHARACTERISTICS:
 (A) LENGTH: 18 base pairs
 (B) TYPE: nucleic acid
 (C) STRANDEDNESS: single
 (D) TOPOLOGY: linear
 (vi) ORIGINAL SOURCE:
 (A) ORGANISM: not provided
 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:11
 CGGATAGCTC GAATATCG 18
 (2) INFORMATION FOR SEQ ID NO: 12:
 (i) SEQUENCE CHARACTERISTICS:
 (A) LENGTH: 17 base pairs
 (B) TYPE: nucleic acid
 (C) STRANDEDNESS: single
 (D) TOPOLOGY: linear
 (vi) ORIGINAL SOURCE:
 (A) ORGANISM: not provided
 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:12
 AACATTGCGT CGTTTGC 17
 (2) INFORMATION FOR SEQ ID NO: 13:
 (i) SEQUENCE CHARACTERISTICS:
 (A) LENGTH: 24 base pairs
 (B) TYPE: nucleic acid
 (C) STRANDEDNESS: single
 (D) TOPOLOGY: linear
 (vi) ORIGINAL SOURCE:
 (A) ORGANISM: not provided
 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:13
 GTAATTACCA GGCAAGGACT CTCC 24
 (2) INFORMATION FOR SEQ ID NO: 14:
 (i) SEQUENCE CHARACTERISTICS:
 (A) LENGTH: 24 base pairs
 (B) TYPE: nucleic acid
 (C) STRANDEDNESS: single
 (D) TOPOLOGY: linear
 (vi) ORIGINAL SOURCE:
 (A) ORGANISM: not provided
 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:14
 GTCATCGGCG GGGGTCATAA CGTG 24
 (2) INFORMATION FOR SEQ ID NO: 15:
 (i) SEQUENCE CHARACTERISTICS:
 (A) LENGTH: 24 base pairs
 (B) TYPE: nucleic acid
 (C) STRANDEDNESS: single
 (D) TOPOLOGY: linear
 (vi) ORIGINAL SOURCE:
 (A) ORGANISM: not provided
 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:15
 GAGGATAGAG AACACTACGA ATCG 24
 (2) INFORMATION FOR SEQ ID NO: 16:
 (i) SEQUENCE CHARACTERISTICS:
 (A) LENGTH: 20 base pairs
 (B) TYPE: nucleic acid
 (C) STRANDEDNESS: single
 (D) TOPOLOGY: linear
 (vi) ORIGINAL SOURCE:
 (A) ORGANISM: not provided
 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:16
 CAGGTCAAGT CAATAGATTC 20
 (2) INFORMATION FOR SEQ ID NO: 17:
 (i) SEQUENCE CHARACTERISTICS:
 (A) LENGTH: 22 base pairs
 (B) TYPE: nucleic acid
 (C) STRANDEDNESS: single
 (D) TOPOLOGY: linear
 (vi) ORIGINAL SOURCE:
 (A) ORGANISM: not provided
 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:17
 CAGAATTTAG TGAATAATAT TG 22
 (2) INFORMATION FOR SEQ ID NO: 18:
 (i) SEQUENCE CHARACTERISTICS:
 (A) LENGTH: 20 base pairs
 (B) TYPE: nucleic acid
 (C) STRANDEDNESS: single
 (D) TOPOLOGY: linear
 (vi) ORIGINAL SOURCE:
 (A) ORGANISM: not provided
 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:18
 GCCAGAGATA ATGCTATTCC 20
 (2) INFORMATION FOR SEQ ID NO: 19:
 (i) SEQUENCE CHARACTERISTICS:
 (A) LENGTH: 23 base pairs
 (B) TYPE: nucleic acid
 (C) STRANDEDNESS: single
 (D) TOPOLOGY: linear
 (vi) ORIGINAL SOURCE:
 (A) ORGANISM: not provided
 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:19
 CATTGATCCA TATATGACAT CAC 23
 (2) INFORMATION FOR SEQ ID NO: 20:
 (i) SEQUENCE CHARACTERISTICS:
 (A) LENGTH: 41 base pairs
 (B) TYPE: nucleic acid
 (C) STRANDEDNESS: single
 (D) TOPOLOGY: linear
 (vi) ORIGINAL SOURCE:
 (A) ORGANISM: not provided
 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:20
 GTGATGTCAT ATATGGATCA ATGGGAAGAG GTAGGGTTCA G 41
 (2) INFORMATION FOR SEQ ID NO: 21:
 (i) SEQUENCE CHARACTERISTICS:
 (A) LENGTH: 21 base pairs
 (B) TYPE: nucleic acid
 (C) STRANDEDNESS: single
 (D) TOPOLOGY: linear
 (vi) ORIGINAL SOURCE:
 (A) ORGANISM: not provided
 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:21
 CAAGAGTCGG TGGAATATTC G 21
 (2) INFORMATION FOR SEQ ID NO: 22:
 (i) SEQUENCE CHARACTERISTICS:
 (A) LENGTH: 41 base pairs
 (B) TYPE: nucleic acid
 (C) STRANDEDNESS: single
 (D) TOPOLOGY: linear
 (vi) ORIGINAL SOURCE:
 (A) ORGANISM: not provided
 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:22
 CGAATATTCC ACCGACTCTT GGTACGCTTC TCCTACTCTA T 41
 (2) INFORMATION FOR SEQ ID NO: 23:
 (i) SEQUENCE CHARACTERISTICS:
 (A) LENGTH: 21 base pairs
 (B) TYPE: nucleic acid
 (C) STRANDEDNESS: single
 (D) TOPOLOGY: linear
 (vi) ORIGINAL SOURCE:
 (A) ORGANISM: not provided
 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:23
 CTAATAAGTA AGATCGCGGA A 21
 (2) INFORMATION FOR SEQ ID NO: 24:
 (i) SEQUENCE CHARACTERISTICS:
 (A) LENGTH: 41 base pairs
 (B) TYPE: nucleic acid
 (C) STRANDEDNESS: single
 (D) TOPOLOGY: linear
 (vi) ORIGINAL SOURCE:
 (A) ORGANISM: not provided
 (xi) SEQUENCE DESCRIPTION: SEQ ID NO:24
 TTCCGCGATC TTACTTATTA GCATGGAGAG GATACTTGAA C 41