bluuebunny/crossref_metadata_2025 huggingface dataset:
Internal structure of mixed‐sand‐and‐gravel beach deposits revealed using ground‐penetrating radar
Authors: Neal Adrian , Pontee Nigel I , Pye Ken , Richards Julie
Published: March 2003

ABSTRACT Mixed‐sand‐and‐gravel beaches are a distinctive type of coarse‐clastic beach. Ground‐penetrating radar (GPR) and photographic records of previous excavations are used to investigate the stratigraphy and internal sedimentary structure of mixed‐beach deposits at Aldeburgh in Suffolk, south‐east England. The principles of radar stratigraphy are used to describe and interpret migrated radar reflection profiles obtained from the study site. The application of radar stratigraphy allows the delineation of both bounding surfaces (radar surfaces) and the intervening beds or bed sets (radar facies). The deposits of the main backshore berm ridge consist of seaward‐dipping bounding surfaces that are gently onlapped by seaward‐dipping bed sets. Good correspondence is observed between a sequence of beach profiles, which record development of the berm ridge on the backshore, and the berm ridge's internal structure. The beach‐profile data also indicate that backshore berm ridges at Aldeburgh owe their origin to discrete depositional episodes related to storm‐wave activity. Beach‐ridge plain deposits at the study site consist of a complex, progradational sequence of foreshore, berm‐ridge, overtop and overwash deposits. Relict berm‐ridge deposits, separated by seaward‐dipping bounding surfaces, form the main depositional element beneath the beach‐ridge plain. However, the beach ridges themselves are formed predominantly of vertically stacked overtop/overwash units, which lie above the berm‐ridge deposits. Consequently, beach‐ridge development in this progradational, mixed‐beach setting must have occurred when conditions favoured overtopping and overwashing of the upper beachface. Interannual to decadal variations in wave climate, antecedent beach morphology, shoreline progradation rate and sea level are identified as the likely controlling factors in the development of such suitable conditions.
Phylogeny of New Zealand hepialid moths (Lepidoptera: Hepialidae) inferred from a cladistic analysis of morphological data
Authors: Brown B. , Dugdale J. S. , Emberson R. M. , Paterson A. M.
Published: March 2003

The phylogeny of the New Zealand hepialid moths was estimated from a cladistic analysis of sixty‐three morphological characters, from all life cycle stages. One hundred and sixteen maximum parsimony trees were produced. The phylogenetic reconstruction indicated that the currently recognized generic concepts, and the four informal lineages hypothesized in a previous morphological taxonomic revision, were monophyletic. The relationships of species within genus Wiseana were not fully resolved. Analysis of a data set of thirty‐nine adult male characters from the New Zealand taxa and the Australian genera Jeana, Oxycanus and Trictena supported the monophyly of the New Zealand ‘Oxycanus’ s.s lineage.
Phylogeny of the subfamilies of Chironomidae (Diptera)
Authors: Sæther O. A.
Published: March 2003

Summary The phylogeny of the subfamilies of Chironomidae are cladistically analysed using parsimony. A data matrix is presented and some characters discussed. Different outgroup taxa, constraints and options are used, characters unordered or ordered, weighted or unweighted, the results reweighted or not and the results discussed. Telmatogetoninae in all cladograms forms the sister group of the remaining subfamilies. Aphroteniinae in some cladograms forms the sister group of all subfamilies except Telmatogetoninae, whereas in other cladograms, including the preferred cladogram, it is part of Tanypodoinae, which otherwise includes Podonominae, Usumbaromyiinae and Tanypodinae. Chilenomyiinae is basal in Tanypodoinae in some cladograms. In most cladograms, including the preferred cladogram, it is basal in Chironomoinae, which also includes Buchonomyiinae, Diamesinae, Prodiamesinae, Orthocladiinae and Chironominae. The preferred cladogram is compared with the relationships between different subfamilies suggested by previous authors.
Restricted rifting and its coexistence with compressional structures: results from the CROP 3 traverse (Northern Apennines, Italy)
Authors: Liotta  , Cernobori  , Nicolicl
Published: March 2003

A geological interpretation of the deep seismic reflection line CROP 3 (Italian program CROsta Profonda: deep crust, profile no. 3), which crosscuts the Northern Apennines from the Tyhrrenian to the Adriatic coast, is presented. The profile images the lithosphere up to 15 s TWT and highlights several peculiarities: (a) the lower crust is bedded by discontinuous sub parallel reflective markers which terminate at ≈ 7 s beneath the western side of the profile; (b) in the same area a notable reflection is recognizable at 10 s and is interpreted as the top of the asthenosphere; (c) east‐plunging shear zones are recognizable throughout the crust. By contrast, the Adriatic (outer) side of this line shows: (a) reflections deepening westward, from 13 to 15 s TWT, related to the base of the crust (33–38 km depth); (b) the absence of thick bedding of reflective markers within the lower crust; and (c) tectonic structures affecting the basement which are different from those which deform the cover of the Northern Apennines. Geological interpretation is based on the eastward migration of ductile shear zones which have been recognized on the seismic line. The bending of the outer zone crust is considered to be a consequence of the rifting process with the application of pushing forces against the Adriatic lithosphere which cannot escape toward east.
Recognizable primary volcanic and sedimentary features in a low‐strain domain of the highly deformed, oldest known (≈ 3.7–3.8 Gyr) Greenstone Belt, Isua, West Greenland
Authors: Appel  , Fedo  , Moorbath  , Myers
Published: March 2003

A low‐strain domain has been identified in the metamorphosed, mostly highly deformed volcanic and sedimentary rocks of the early Archaean Isua supracrustal belt. This domain contains well‐preserved volcanic and sedimentary features, including basaltic pillow lavas, pillow breccia, heterogeneous volcanic breccia, amygdules in metabasalt, and polymict conglomerate dominated by recrystallized chert and volcanic clasts. The low‐strain domain is bounded by highly deformed rocks mostly derived from basalt, chert, and banded iron formation. These discoveries demonstrate that some primary features have escaped the pervasive metasomatism dominant in other parts of the belt and, furthermore, strengthen the characterization of the Isua supracrustals as a typical greenstone belt.
Ultra‐high pressure (P > 6 GPa) garnet peridotites in Western Norway: exhumation of mantle rocks from > 185 km depth
Authors: Van Roermund  , Drury
Published: March 2003

We report here for the first time the occurrence of relics of majoritic garnet within orogenic garnet peridotites from Otrøy, Western Gneiss Region, Norway. The microstructural evidence consists of two‐pyroxene exsolution from garnet. Majoritic garnets are only stable at depths greater than 150 km. Estimates of the initial composition of the majoritic garnets imply pressures of 6–6.5 GPa indicating that the Otrøy peridotites were derived from depths > 185 km.

 Mineral‐chemical data indicate that the present mineral compositions equilibrated at mantle conditions around 805 ± 40 °C and 3.2 ± 0.2 GPa.

 Estimates of the initial pressure temperature (PT) conditions and PTtime (t) path are consistent with a multistage, multiorogenic exhumation history with upwelling of hot asthenosphere up to ≈ 100 km in the Pre‐Cambrian followed by subsequent crustal emplacement and exhumation during the Caledonian orogeny.
The transition from rifting to sea‐floor spreading within a magma‐poor rifted margin: field and isotopic constraints
Authors: Schaltegger Urs , Desmurs Laurent , Manatschal Gianreto , Müntener Othmar , Meier Martin , Frank Martin , Bernoulli Daniel
Published: March 2003

We provide new geological and isotope geochemical constraints on the evolution from continental rifting to sea‐floor spreading along a segment of the Jurassic Tethyan margin exposed in the Platta and Err nappes (eastern Central Alps). Field observations show that the ocean–continent transition zone is characterized by oceanward‐dipping detachment faults leading to the exhumation of subcontinental mantle rocks subsequently intruded by gabbro bodies and dolerite dikes, and covered by pillow basalts and radiolarites. Zircons extracted from gabbros and albitite yield concordant U–Pb ages of 161 ± 1 Ma; their initial ɛHf (+ 14.4 to + 14.9) as well as bulk rock ɛNd values of from gabbros and basalts (+ 7.3 to + 9.5) point to a MORB‐type depleted mantle source. These data suggest that the onset of magmatic activity coincides with the latest phase of mantle exhumation along low‐angle detachment faults and may be controlled by upwelling asthenosphere beneath a zone of exhumed continental mantle.
Astronomical tuning of the Tortonian 87Sr/86Sr curve in the Mediterranean basin
Authors: Sprovieri Mario , Barbieri Mario , Bellanca Adriana , Neri Rodolfo
Published: March 2003

ABSTRACT This work presents a detailed 87Sr/86Sr isotope curve for the interval 7.5–9.7 Ma obtained by a high‐resolution analysis (sampling spacing of about 40 kyr) of an astronomically calibrated land‐based sedimentary sequence exposed in the central Mediterranean area (Gibliscemi section, southern Sicily). The main aim is to verify a synchronous response of the Mediterranean seawater Sr isotope record to the oceanic forcing on the basis of multiple comparisons of the Gibliscemi record with published coeval 87Sr/86Sr curves. A good correlation with the 87Sr/86Sr data from the ODP site 926 (equatorial Atlantic ocean), considered to be the Sr chemostratigraphic reference section for the Late Miocene, and from the Pacific DSDP site 590B was registered. Conversely, the comparison of the Gibliscemi Sr isotope data with 87Sr/86Sr ratios from the coeval segment of the land‐based Sardella section (eastern Mediterranean) shows important differences highlighting a local control on the seawater Sr isotope changes in semi‐isolated subbasins within the Late Miocene Mediterranean.
PCR‐aided genomic sequencing of 5' subterminal sequences of the maize transposable element Activator (Ac) in transgenic tobacco plants.
Authors: Ott T , Nelsen‐Salz B , Doring HP
Published: March 2003

The maize transposable element Activator (Ac) carries subterminal CpG‐ rich sequences which are essential for the transposition of the element. It has previously been shown that the methylation of certain sequences contained in this region can alter their ability to interact with the Ac‐encoded protein. The novel hypothesis that the methylation of subterminal Ac sequences is required for transposition was tested. Approximately 150 bp of the 5' subterminal region of the Ac element was examined for the presence of 5‐methylcytosines by the ligation‐mediated polymerase chain reaction (LMPCR)‐aided genomic sequencing method. The methylation status of 22 and 39 cytosines on either strand of the DNA were analysed in each of five different transgenic tobacco cultures carrying transposable Ac sequences. Ten micrograms of tobacco DNA were used for each base‐specific cleavage reaction before amplification by LMPCR. All but one of the cytosines were unmethylated. Only a minor fraction of the Ac molecules was methylated at one cytosine residue. It is concluded that DNA methylation at the tested Ac sequences is not required for the transposability of Ac or Ds elements in tobacco cells.
Potato virus X as a vector for gene expression in plants
Authors: Chapman Sean , Kavanagh Tony , Baulcombe David
Published: March 2003

The suitability of potato virus X (PVX) as a gene vector in plants was tested by analysis of two viral constructs. In the first, the GUS gene of Escherichia coli was substituted for the viral coat protein gene. In the second, GUS was added into the viral genome coupled to a duplicated copy of the viral promoter for the coat protein mRNA. The viral construct with the substituted coat protein gene accumulated poorly in inoculated protoplasts and failed to spread from the site of infection in plants. These results suggest a role for the viral coat protein in key stages of the viral infection cycle and show that gene replacement constructs are not suitable for the production of PVX‐based gene vector. The construct with GUS coupled to the duplicated promoter for coat protein mRNA also accumulated less well in protoplasts than the unmodified PVX, but did infect systemically and directed high level synthesis of GUS in inoculated and systemically infected tissue. Although there was some genome instability in the PVX construct, much of the viral RNA in the systemically infected tissue had retained the foreign gene insertion, especially in infected Nicotiana clevelandii plants. These data point to a general utility of PVX as a vector for unregulated gene expression in plants.
Cis‐analysis of a seed protein gene promoter: the conservative RY repeat CATGCATG within the legumin box is essential for tissue‐specific expression of a legumin gene
Authors: Bäumlein Helmut , Nagy† Istvan , Villarroel Raimundo , Inzé Dirk , Wobus Ulrich
Published: March 2003

A 2.4 kb fragment containing the 5 ‐flanking region and the 5′‐noncoding sequence of the Vicia faba legumin gene LeB4 mediates high level seed‐specific expression in transgenic tobacco plants. Deleted derivatives of this legumin upstream sequence were fused to the npt‐ll reporter gene to determine the tissue‐specific activity of the chimeric constructs in stably transformed tobacco plants. The results indicate the presence of positive regulatory, enhancer‐like cis elements within 566 bp of the upstream sequence. Most importantly, however, these elements are only fully functional in conjunction with the core motif CATGCATG of the legumin box around position ‐95, since destruction of the motif by a 6 bp deletion in an otherwise intact 2.4 kb upstream sequence drastically reduces expression in seeds. At the same time, low level expression in leaves is observed. The occurrence of similar CATGCATG consensus cis elements with alternating purine and pyrimidine base pairs in front of several other plant genes suggests a functional role of the motif in a wider range of plant promoters.
Identification of an S‐locus glycoprotein allele introgressed from B. napus ssp. rapifera to B. napus ssp. oleifera
Authors: Goring Daphne R. , Banks Paul , Fallis Lynne , Baszczynski Chris L. , Beversdorf Wallace D. , Rothstein Steven J.
Published: March 2003

Self‐incompatible Brassica napus ssp. oleifera lines were generated by introgressing the S‐locus from the self‐incompatible B. napus ssp. rapifera Z line into the self‐compatible cultivars, Topas and Regent, resulting in T2 and R2, respectively. Screening of a cDNA library made from R2 stigma RNA produced several candidate SLG (S‐locus glycoprotein) cDNAs. One of the cDNAs, A14, was found to be represented in only the R2, T2 and Z lines. In addition, the corresponding A14 gene was demonstrated to segregate with the T2 self‐incompatibility phenotype in an F2 population derived from a cross between T2 and Topas, and to exhibit high mRNA levels in the stigmas prior to anthesis. Sequence analysis of the A14 cDNA revealed close homology to B. oleracea SLG alleles associated with a Class I high activity self‐incompatibility phenotype.
The plant homologue of MAP kinase is expressed in a cell cycle‐dependent and organ‐specific manner
Authors: Jonak Claudia , Páy Aniko , Börge Laszlo , Hirt Heribert , Heberle‐Bors Erwin
Published: March 2003

In animals, MAP kinase plays a key role in growth factor‐stimulated signalling and in mitosis. The isolation of a Medicago sativa cDNA clone MsK7 which shows 52% identity to animal MAP kinases is reported. The deduced protein sequence shows all the important structural features of MAP kinases and also contains the highly conserved Thr‐183 and Tyr‐185 residues. Northern analysis of synchronized alfalfa cells showed that the MsK7 kinase gene is expressed at low levels in G1 phase but at higher levels in S and G2 phases of the cell cycle. In the plant, only stems and roots were found to contain MAP kinase MsK7 mRNA. Southern and PCR analyses indicated that alfalfa contains at least four highly related MAP kinase genes.
A 42 bp promoter fragment of the gene for subunit III of photosystem I (psaF) is crucial for its activity
Authors: Flieger K. , Tyagi A. , Sopory S. , Cseplö A. , Herrmann R.G. , Oelmüller R.
Published: March 2003

The promoter of the gene for the subunit III of photosystem I reaction center (psaF) from spinach has been dissected and studied via promoter/GUS gene fusions in transgenic tobacco. It possesses an architecture that differs from any other spinach promoter of genes encoding proteins involved in photosynthesis studied to date. A 42 bp region located between −220 and −179 bp upstream of the transcription start site has been identified that is indispensable for expression and binds a trans‐acting factor. Maximal light‐response is obtained with a −220/+163 bp segment, whereas longer promoter sequences are significantly less effective, indicating the existence of upstream elements with silencer characteristics. F1 seedlings show different spatial expression patterns in darkness or light. Etiolated seedlings display high GUS activity in the upper hypocotyl, the hook region and the vascular tissue of the cotyledons, whereas in light‐grown seedlings no activity was detected in the hypocotyl and almost all cells of the cotyledons express the GUS gene.
Soluble isoforms of starch synthase and starch‐branching enzyme also occur within starch granules in developing pea embryos
Authors: Denyer Kay , Sidebottom Christopher , Hylton Christopher M. , Smith Alison M.
Published: March 2003

Developing wild‐type pea embryos contain two major isoforms of starch synthase and two isoforms of starch‐branching enzyme. One of the starch synthases and both starch‐branching enzymes occur both in the soluble fraction and tightly bound to starch granules. The other starch synthase, which is very similar to the waxy proteins of other species, is exclusively granule‐bound. It is inactive when solubilized in a native form from starch granules, but activity is recovered when the SDS‐denatured protein is reconstituted from polyacrylamide gels.

Evidence is presented which indicates that all of these proteins become incorporated within the structure of the granule as it grows. It is proposed that the granule‐bound waxy protein is active in vivo at the granule surface, whereas the remaining proteins are active in the soluble fraction of the amyloplast. The proteins become trapped within the granule matrix as the polymers they synthesize crystallize around them, and they probably play no further part in polymer synthesis.
Oligogalacturonides inhibit the formation of roots on tobacco explants
Authors: Bellincampi Daniela , Salvi Giovanni , De Lorenzo Giulia , Cervone Felice , Marfà Victòria , Eberhard Stefan , Darvill Alan , Albersheim Peter
Published: March 2003

α‐1,4‐Oligogalacturonides with degrees of polymerization (DPs) ranging from 6 to 18 or 2 to 8 were added to tobacco leaf explants and root formation was evaluated after 15 days of incubation. Auxin‐induced formation of roots was inhibited by oligogalacturonides with DPs 6–18 but not by the oligogalacturonides with DPs 2–8. The inhibition of root formation by the larger oligogalacturonides was prevented by increasing the amount of auxin present in the medium. Oligogalacturonides (DPs 6–18) also inhibited root formation when added to tobacco thin cell‐layer (TCL) explants in a medium that is known to induce the formation of roots. The addition of size‐homogeneous oligogalacturonides, to either tobacco leaf explants or TCLs, established that oligogalacturonides with DPs between 10 and 14 were most active in inhibiting the formation of roots. These data suggest that oligogalacturonides of the same size as those known to elicit plant defense responses, and to affect floral development and membrane functions, also inhibit the induction of root morphogenesis in tobacco.
Functional expression of the Erwinia uredovora carotenoid biosynthesis gene crtl in transgenic plants showing an increase of β‐carotene biosynthesis activity and resistance to the bleaching herbicide norflurazon
Authors: Misawa Norihiko , Yamano Shigeyuki , Linden Hartmut , de Felipe Maria R. , Lucas Mercedes , Ikenaga Hiroshi , Sandmann Gerhard
Published: March 2003

Among the enzymes involved in carotenoid biosynthesis, phytoene desaturase is considered to be a rate‐limiting enzyme in this pathway and is also the target of many bleaching herbicides. This enzyme shows diversity concerning its function and amino acid homology among various organisms. The phytoene desaturase gene crtl of Erwinia uredovora was expressed, the 5′‐region of which was fused to the sequence for the transit peptide of a pea Rubisco small subunit, in tobacco plants under the control of the CaMV 35S promoter. This chimeric gene product was targeted into chloroplasts and processed in the transgenic plants. The production and processing of the corresponding protein could be demonstrated by Western blotting. Immunogold localization showed that the location of the gene product Crtl was preferentially in the thylakoids. A radioactive labeling study using the leaves demonstrated enhanced activity for the synthesis of β‐carotene. In addition, the transgenic tobacco acquired elevated resistance to the bleaching herbicide norflurazon.
Cloning and expression of flavonol synthase from Petunia hybrida
Authors: Holton Timothy A. , Brugliera Filippa , Tanaka Yoshikazu
Published: March 2003

Flavonols are important co‐pigments in flower colour and are also essential for pollen tube growth. In petunia, flavonol synthesis is controlled by the Fl locus. Flavonol synthase (FLS) belongs to the 2‐oxoglutarate‐dependent dioxygenase family. Dioxygenase gene fragments were amplified by PCR on cDNA made from FlFl and flfl flowers using degenerate primers designed from conserved dioxygenase sequences. A petunia petal cDNA library was screened for clones that hybridized more strongly to the Fl PCR products than the fl PCR products. A full‐length cDNA clone identified by this screening exhibited FLS activity when expressed in yeast. FLS gene expression is developmentally regulated during flower development. Antisense expression of an FLS cDNA clone in petunia markedly reduced flavonol synthesis in petals. RFLP mapping showed that the FLS gene is linked to Fl, suggesting that Fl is the structural gene for FLS.
Characterization of cDNAs for stylar transmitting tissue‐specific proline‐rich proteins in tobacco
Authors: Cheung Alice Y. , May Bruce , Kawata Evelynn E. , Gu Qing , Wu Hen‐ming
Published: March 2003

The pistil of flowers is a specialized organ which contains the female gametophytes and provides the structures necessary for pollination and fertilization. Pollen deposited on the stigmatic surface of a compatible plant germinates a pollen tube which penetrates the stigmatic papillae and grows intercellularly through the style towards the ovules in the ovary. Pollen tube growth is largely restricted to the transmitting tissue in the style. Therefore the stylar transmitting tissue is extremely important for the migration of the pollen cell towards the ovary. We have isolated two related cDNAs, transmitting tissue‐specific (TTS)‐1 and TTS‐2, derived from two proline‐rich protein (PRP)‐encoding mRNAs that accumulate specifically in the transmitting tissue of tobacco. The deduced PRP sequences share similarities with proline‐rich cell wall glycoproteins found in a variety of plants. TTS‐1 and TTS‐2 mRNAs are induced in very young floral buds, accumulate most abundantly during the later stages of flower development when style elongation is the most rapid, and remain at relatively high levels at anthesis. These mRNAs become undetectable in maturing green fruits. In situ hybridization shows that TTS‐1 and TTS‐2 mRNA accumulation is restricted to the transmitting tissue of the style. The possible roles that these transmitting tissue‐specific PRPs may play in maintaining the structural integrity of the style or in the function of this organ is discussed.
The β‐phaseolin gene is flanked by matrix attachment regions
Authors: van der Geest Apolonia H.M. , Hall Gerald E. , Spiker Steven , Hall Timothy C.
Published: March 2003

The β‐phaseolin promoter yields high levels of spatially and temporally regulated expression in transgenic plants. It is shown that elements within 5′ and 3′ regions flanking the coding sequence bind strongly and specifically to matrix preparations from tobacco nuclei. Both of these matrix attachment regions (MARs) are A/T‐rich and contain topoisomerase II, Abox, T‐box and MAR motifs frequently found in MARs in Drosophila and other organisms. The β‐phaseolin 5′ MAR is located on a 1047 bp fragment with a core element of 673 bp (positions −1096 to −424), that includes a 55 bp region (−682 to −628) previously shown to function as an enhancer. A 1205 bp fragment (positions +1879 to +3083) 3′ of the phaseolin coding region also showed strong and specific binding to nuclear matrix preparations and contained consensus MAR motifs. A β‐phaseolin promoter‐GUS fusion construct (p1470/phas) that included the MAR elements exhibited higher expression levels and lower plant‐to‐plant variability in stably transformed tobacco plants than did a similar construct lacking MAR sequences. Seeds of plants containing two intact copies of the insert had higher expression levels than did those of plants containing a single copy. These findings, which are the first to report a plant gene being flanked both 5′ and 3′ by MARs, are in accordance with previous observations of increased DNase I sensitivity associated with the onset of phaseolin transcription in bean and with the consistently high and position‐independent expression observed in transgenic plants from the β‐phaseolin promoter. The results suggest that the native β‐phaseolin gene is located on a small chromatin loop that functions as an active and independent domain.
Cloning and structural analysis of the anthocyanin pigmentation locus Rt of Petunia hybrida: characterization of insertion sequences in two mutant alleles
Authors: Kroon Johan , Souer Erik , De Graaff Alja , Xue Yongbiao , Mol Joseph , Koes Ronald
Published: March 2003

Anthocyanin biosynthesis in flowers of Petunia hybrida is controlled by the regulatory genes an1, an2 and an11. Seven classes of cDNA clones homologous to transcripts that are down‐regulated in an1−, an2− and an11− mutants were isolated via differential cDNA cloning. Genetic mapping, antisense RNA experiments and analyses of mutant alleles demonstrated that one class of clones originated from the Rt locus. The rt gene has no introns and encodes a protein with homology to mammalian glucuronosyl transferases and flavonoid 3‐O‐glucosyl‐transferase (UF3GT) encoded by the bz1 gene from Zea mays. As the Rt locus controls the rhamnosylation of reddish anthocyanin‐3‐O‐glucosides which is the first in a series of modifications that finally yield magenta or blue/purple coloured anthocyanins, this suggests that rt encodes an anthocyanin rhamnosyl transferase.

Molecular analysis of two mutant rt alleles showed that their expression is blocked by different DNA insertion elements. Mutability of the rt‐vu15 allele results from the presence of a 284 bp transposable element (dTph1) in the rt promoter region, causing a block in transcription. The protein coding region of the rt‐r27 allele contains a 442 bp insertion (dTph3) resulting in premature polyadenylation of rt transcripts. Although dTph3 cannot transpose, it has sequence characteristics of transposable elements, suggesting that it is a defective member of a new family of transposable elements.
Mutations at two new Arabidopsis ABA response loci are similar to the abi3 mutations
Authors: Finkelstein Ruth R.
Published: March 2003

Two new Arabidopsis loci involved in ABA response, ABA‐insensitive (ABI)4 and ABI5, have been identified by mutation. The abi4 and abi5 mutants were characterized in terms of ABA sensitivity of seed germination, dormancy, seed‐specific gene expression and stomatal regulation. Their phenotypes are similar to mutations in ABI3, which is thought to encode a seed‐specific transcriptional activator. Analysis of double mutants combining abi4 and abi5 mutations with abi1, abi2 or abi3 mutations also suggests that ABI4 and ABI5 act in the same transduction pathway as ABI3.
Changes in pectin structure and localization during the growth of unadapted and NaCl‐adapted tobacco cells
Authors: McCann Maureen C. , Shi Jun , Roberts Keith , Carpita Nicholas C.
Published: March 2003

Tobacco cells adapted to grow in high concentrations of NaCl exhibit a drastically altered growth physiology that results in cells whose fully expanded volume is only one‐fifth to one‐eighth those of unadapted cells. Comparison between NaCl‐adapted and unadapted tobacco cells provides an opportunity to evaluate current concepts of the structural and mechanical determinants of cell wall expansion. Both biochemical studies of pectic polymers and the ultrastructural localization of pectic epitopes at three specific phases of cell culture, maximal cell division, maximal elongation, and stationary phase are reported here.

One‐half of the galactosyluronic acid units in wall polymers of NaCl‐adapted cells are esterified throughout the culture period, while wall polymers of unadapted cells show a rise in esterified polygalacturonic acid from 50 to 80% during elongation and then a decrease to 70% at stationary phase. Methyl esters account for only a proportion of the total esterified polygalacturonic acid at any stage in both unadapted and NaCl‐adapted cell walls. Using monoclonal antibodies, we show differences in the localization of relatively methyl‐esterified and unesterified pectic epitopes at different stages of growth and corroborate the chemical determinations. Fourier transform infrared (FTIR) microspectroscopy of representative walls of both NaCl‐adapted and unadapted cells confirms, at the single cell wall level, that results obtained from chemical analysis of bulk samples are applicable to the entire cell population.

FTIR microspectroscopy also reveals an increase in wall protein in the walls of adapted cells. Images obtained by the fast‐freeze, deep‐etch, rotary‐shadowed replica technique show clearly different cell wall architectures in NaCl‐adapted compared with unadapted cells; walls of elongating unadapted cells contain long, thin fibres that show a net orientation with respect to the long axis of the cell, whereas walls of adapted cells have thicker, flatter bundles of fibres with no clear net orientation. Polarized FTIR microspectroscopy indicates that, in unadapted tobacco cells during elongation, pectin molecules may be oriented within the wall in a similar manner to cellulose. Possible ways in which pectin structure and conformation may affect the behaviour of the cellulose‐xyloglucan network are discussed.
Identification of S‐locus linked PCR fragments in rye (Secale cereale L.) by denaturing gradient gel electrophoresis
Authors: Wehling P. , Hackauf B. , Wricke G.
Published: March 2003

Rye inbred lines segregating at the S‐locus and homozygous at the Z‐locus were investigated by PCR with primers derived from Brassica SLG‐sequences. After denaturing gradient gel electrophoresis (DGGE), a 280 bp PCR‐fragment displays a polymorphism perfectly correlated to the underlying S‐genotypes. This is the first report on S‐related DNA polymorphism in a bifactorial self‐incompatibility system of the Poaceae.
Water channels in the plant plasma membrane cloned by immunoselection from a mammalian expression system
Authors: Kammerloher Werner , Fischer Utz , Piechottka Gerd P. , Schäffner Anton R.
Published: March 2003

Expression in mammalian COS cells and an efficient microtiter‐based strategy for immunoselection was used in a novel approach to identify genes encoding plant membrane proteins. COS cells were transfected with an Arabidopsis thaliana root cDNA library constructed in a bacterial mammalian shuttle vector and screened with an antiserum raised against purified deglycosylated integral plasma membrane proteins from A. thaliana roots. Antibodies directed against a prominent 27 kDa antigen led to the identification of five different genes. They comprised two subfamilies related to the major intrinsic protein (MIP) superfamily and were named plasma membrane intrinsic proteins, PIP1 and PIP2, since the cellular localization of PIP1 and most probably PIP2 proteins in the plasma membrane was independently confirmed by their co‐segregation with marker enzymes during aequeous two‐phase partitioning. Surprisingly, expression in Xenopus laevis oocytes revealed that all five PIP mRNAs coded for Hg2+‐sensitive water transport facilitating activities. There had been no previous evidence of the existence of water channels in the plasma membrane of plant cells and the high diffusional water permeability of the lipid bilayer was considered to be sufficient for water exchange. Nevertheless, Northern and Western analyses showed that the PIP genes are constitutively and possibly even redundantly expressed from the small A. thaliana genome.
Malate‐sensitive anion channels enable guard cells to sense changes in the ambient CO2 concentration
Authors: Hedrich Rainer , Marten Irene , Lohse Gabi , Dietrich Petra , Winter Heike , Lohaus Gertrud , Heldt Hans‐Walter
Published: March 2003

Malate is a characteristic metabolite in the photosynthesis of C4 and CAM plants. Furthermore, changes in the intracellular concentration of this organic acid provide part of the osmotic motor for guard cells. Since alterations in the malate concentration influence the photosynthetic capacity on one side and stomatal action on the other, it was studied whether the extracellular malate level represents an indicator of changes in the ambient CO2 concentration and a key regulator of ion transport in guard cells.

Here it is demonstrated that alterations in the ambient CO2 level modify the extracellular malate concentration of Vicia faba leaves. Elevated external malate caused stomatal closure in a concentration‐dependent manner (Kmmal = 0.3 mM). Slight variations in the external malate concentration strongly regulate the voltage‐dependent properties of GCAC1, an anion‐release channel in the plasma membrane of guard cells. Superfusion of guard cell protoplasts with malate levels in the physiological range (Kmmal = 0.4 mM) caused the voltage gate to shift towards the resting potential of the cell‐activating GCAC1. Single‐channel conductance was dependent on the extracellular chloride concentration (KmCl = 3 mM). In the absence of extracellular chloride the plasma membrane lacked anion conductance until the addition of malate induced channel opening. Isophthalate was a powerful agonist in both malate‐induced processes, channel regulation and stomatal closure, indicating that modulation of GCAC1 is a key step in stomatal action. It was thus concluded that feedback regulation of volume and turgor with respect to the ambient CO2 concentration via malate‐sensitive anion channels may provide a CO2 sensor to guard cells.
Isolation and immobilization of various plastid subtypes by magnetic immunoabsorption
Authors: Kausch Albert P. , Bruce Barry D.
Published: March 2003

Antibodies have been prepared which immuno‐localize to the outer membrane of the pea chloroplast envelope and cause agglutination of isolated chloroplasts. This antisera is immunoreactive with a variety of plastid forms from both monocotyledonous and dicotyledonous plants. Whether such antibodies might be effectively used for isolation and immobilization of plastids from whole cell lysates has been tested. A system has been developed for immunolabeling various forms of higher plant plastids with biotinylated antibody and streptavidin magnetic nano‐particles followed by separation of the plastids in a 0.6 Tesla high gradient magnetic field. Using this magnetic immunoabsorption procedure it has been possible to achieve a high degree of positive enrichment for chromoplasts, amyloplasts, and chloroplasts from whole cell lysates of several plant species. The integrity of these plastids has been examined by in organellar protein synthesis, 14C‐ADP‐glucose uptake, flow cytometry, in vitro synthesized precursor import and FITC‐cationized ferritin staining of the plastid envelope. Western blot analysis showed significant enrichment for amyloplasts from cytosolic sucrose synthase in maize endosperm. Magnetic immunoabsorption of subcellular structures from whole cell lysates is a new method that may be useful in the in vitro analysis of many different cellular compartments from a wide range of organisms.
Transgene‐mediated suppression of chalcone synthase expression in Petunia hybrida results from an increase in RNA turnover
Authors: Van Blokland R. , Van der Geest N. , Mol J.N.M. , Kooter J.M.
Published: March 2003

Co‐suppression of the pigmentation gene chalcone synthase (chs) in Petunia hybrida by chs transgenes leads to white or variegated flowers and is characterized by a reduction in steady‐state mRNA levels. To determine the level at which suppression occurs different petunia transformants were analysed containing CaMV‐35S RNA promoter‐driven hybrid genes consisting of the β‐glucuronidase gene (uidA) linked to the full‐length chsA cDNA, the 5′ half or to the 3′ half. With these transgenes one out of 12–15 primary transformants showed suppression of the transgenes and of the resident chs genes throughout the flower or in sectors. The reduction in steady‐state chs mRNA was not the result of a transcriptional inactivation event. As determined by nuclear run‐on experiments, transcription of suppressed chs genes was similar to that of non‐suppressed genes. This indicates that co‐suppression occurs post‐transcriptionally. Among individual transformants the transgenes were transcribed at different levels but neither a high nor a low level correlated with a particular degree or pattern of suppression. Surprisingly, even a promoterless chs transgene construct was found to suppress the endogenous chs genes in three out of 15 transformants. It remains, however, unknown whether or not transcription of the transgene locus is required to induce co‐suppression. The data suggest that properties of the chs transgene locus other than the expression level are important for inducing co‐suppression. The possible role of antisense RNA, which was detected in all transformants, ectopic pairing and the structure of the integrated T‐DNAs in the mechanism of the selective increase in chs RNA turnover are discussed.
Spatial and temporal pattern of light‐regulated gene expression during tobacco seedling development: the photosystem II‐related genes Lhcb (Cab) and PsbP (Oee2)
Authors: Kretsch Thomas , Emmler Karlheinz , Schäfer Eberhard
Published: March 2003

The white mustard (Sinapis alba L.) Lhcb1*1 and PsbP*1 genes that code for proteins related to photosystem II (PSII) in chloroplasts were examined by analysis of promoter fragment β‐glucuronidase (GUS) reporter constructs in transgenic tobacco (Nicotiana tabacum L.) seedlings. The endogenous tobacco genes and the introduced mustard genes follow the same kinetics during seedling development and they show the same expression characteristics for light regulation and for the influence of a ‘plastidic signal’. Hence, the cellular environment of the host plant always dominates the regulation of Lhcb1*1 and PsbP*1 gene expression; as with the mustard system clear differences in the temporal pattern and the physiological responses could be seen. The spatio‐temporal pattern of gene expression was analysed in the different organs of the transgenic tobacco seedlings. In the cotyledons, expression at the PsbP*1 promoter starts in advance, and both genes show a rather uniform distribution of expression during seedling development. In the hypocotyl, a sequential basipetal pattern could be detected and a coordinated expression for the two promoters was analysed. The hypocotyl base is only included in this expression pattern if the seedlings receive light at early stages of development, whereas in later stages gene expression is repressed. A model is proposed that divides tobacco seedling development into three main phases.
Specific distribution of mRNAs in maize growing pollen tubes observed by whole‐mount in situ hybridization with non‐radioactive probes
Authors: Torres Miguel‐Angel , Rigau Joan , Puigdomènech Pere , Stiefel Virginia
Published: March 2003

The distribution of a number of specific mRNAs has been observed in maize growing pollen tubes. Whole‐mount in situ hybridization using digoxygenin‐labelled RNA probes has been tested. The technique appears to be a simple, rapid and reliable method in this system, in immature anthers and also in other tissues. Results for three probes are presented. They correspond to a hydroxyproline‐rich glycoprotein (HRGP), an abundant component of the maize cell wall, to an α‐tubulin (encoded by the Tubα 1 gene) highly expressed in the radicular system of the plant and also in pollen, and to an isoform of the malic enzyme, involved in the basic metabolism of the plant. The mRNAs corresponding to these three proteins are differently distributed in the germinating pollen. While HRGP mRNA is only present in the tube, malic enzyme mRNA is only present in the body of the pollen cell, and α‐tubulin mRNA is present in both parts of the cell but shows a higher accumulation in the tip of the pollen tube.
Cellular localization of the Ca2+ binding TCH3 protein of Arabidopsis
Authors: Antosiewicz Danuta M. , Polisensky Diana H. , Braam Janet
Published: March 2003

TCH3 is an Arabidopsistouch (TCH) gene isolated as a result of its strong and rapid upregulation in response to mechanical stimuli, such as touch and wind. TCH3 encodes an unusual calcium ion‐binding protein that is closely related to calmodulin but has the potential to bind six calcium ions. Here it is shown that TCH3 shows a restricted pattern of accumulation during Arabidopsis vegetative development. These data provide insight into the endogenous signals that may regulate TCH3 expression and the sites of TCH3 action. TCH3 is abundant in the shoot apical meristem, vascular tissue, the root columella and pericycle cells that give rise to lateral roots. In addition, TCH3 accumulation in cells of developing shoots and roots closely correlates with the process of cellular expansion. Following wind stimulation, TCH3 becomes more abundant in specific regions including the branchpoints of leaf primordia and stipules, pith parenchyma, and the vascular tissue. The consequences of TCH3 upregulation by wind are therefore spatially restricted and TCH3 may function at these sites to modify cell or tissue characteristics following mechanical stimulation. Because TCH3 accumulates specifically in cells and tissues that are thought to be under the influence of auxin, auxin levels may regulate TCH3 expression during development. TCH3 is upregulated in response to low levels of exogenous indole‐3‐acetic acid (IAA), but not by inactive auxin‐related compounds. These results suggest that TCH3 protein may play roles in mediating physiological responses to auxin and mechanical environmental stimuli.
Construction of an equalized cDNA library from Arabidopsis thaliana
Authors: Kohchi Takayuki , Fujishige Kotomi , Ohyama Kanji
Published: March 2003

Using a kinetic approach, a cDNA library composed of almost equal representations of all genes expressed in the aerial parts of 2‐week‐old Arabidopsis was constructed. A cDNA was synthesized with an oligo dT primer containing a Notl site. A linker containing the nucleotide sequence of Sse8387I which recognizes octanucleotides was added at the ends of the synthesized cDNA. The cDNA was amplified by the polymerase chain reaction (PCR), denatured, and reassociated under modified conditions. Thereafter, the remaining single‐stranded DNA was converted to double‐stranded DNA and amplified by PCR. These equalization steps were repeated three times and the products were cloned unidirectionally into a plasmid vector. Equalization was evaluated by colony hybridization and DNA sequencing. This approach will be applicable to construct a cDNA library suitable for subtraction, differential screening, and expression screening, especially for mRNA species present at very low concentrations in a few cells of a specific tissue.
Characterization of the S locus genes, SLG and SRK, of the Brassica S3 haplotype: identification of a membrane‐localized protein encoded by the S locus receptor kinase gene
Authors: Delorme Valérie , Giranton Jean‐Loïc , Hatzfeld Yves , Friry Aline , Heizmann Philippe , Ariza María José , Dumas Christian , Gaude Thierry , Cock J. Mark
Published: March 2003

The S locus, which controls the self‐incompatibility response in Brassica, has been shown to contain at least two genes. SLG encodes a secreted S locus glycoprotein whilst SRK encodes a putative S locus receptor kinase. SRK has been shown potentially to encode a functional kinase and genetic evidence indicates that this gene is essential for the self‐incompatibility response. Here the characterization of the SRK and SLG genes of a Brassica line homozygous for the S3 haplotype is described. A 120 kDa glycoprotein was identified in stigmas and several lines of evidence indicated that this protein is encoded by the SRK3 gene. First, the 120 kDa glycoprotein was recognized by antibodies raised against peptides based on the SRK3 gene sequence. Secondly, this protein is polymorphic and, in an F2 population segregating for the S3 haplotype, was expressed only in plants possessing the S3 haplotype. Thirdly, the 120 kDa protein was expressed specifically in stigmas. Finally, the 120 kDa protein was only extracted from stigmas in the presence of detergent indicating that it is anchored in the membrane. SRK has been predicted to encode a transmembrane glycoprotein based on the deduced amino acid sequence. Located on the membrane, SRK is in a position to interface between an extracellular recognition event between pollen and pistil and an intracellular signal transduction pathway which initiates the self‐incompatibility response.
Construction of a rice bacterial artificial chromosome library and identification of clones linked to the Xa‐21 disease resistance locus
Authors: Wang Guo‐Liang , Holsten Thomas E. , Song Wen‐Yuan , Wang He‐Ping , Ronald Pamela C.
Published: March 2003

A bacterial artificial chromosome (BAC) library consisting of 11 000 clones with an average DNA insert size of 125 kb was constructed from rice nuclear DNA. The BAC clones were stable in E. coli after 100 generations of serial growth. Transformation of the BAC clones by electroporation into E. coli was highly efficient and increased with decreasing size of the DNA inserts. The library was evaluated for the presence of organellar, repeated, and telomeric sequences. A very low percentage (<0.3%) of the library consisted of chloroplast and mitochondrial clones. Eighteen BACs were identified that hybridized with an Arabidopsis telomere repeat. Sixteen BACs hybridized with the AA genome‐specific repetitive sequence pOs48. Twelve clones were isolated that hybridized with three DNA markers linked to the Xa‐21 disease resistance locus. The results indicate that the BAC system can be used to clone and manipulate large pieces of plant DNA efficiently.