Abstract:
The present invention relates to transgenic sugar beet plants which due to the expression of cp4/epsps enzyme activity tolerate treatment with about 4 to about 18 liters Roundup® per hectar. The plants can be characterized by theis specific integration site. The invention further relates to seeds obtained from said plants and a method for producing said plants.

Description:
CROSS-REFERENCE TO RELATED APPLICATION 
     U.S. application Ser. No. 09/434,039 is a continuation of U.S. application Ser. No. 09/182,117 filed Oct. 29, 1998, which has issued as U.S. Pat. No. 6,204,436, which claims the benefit of provisional application No. 60/112,003 filed Oct. 31, 1997. 
    
    
     BACKGROUND OF THE INVENTION 
     1. Field of the Invention 
     The present invention relates to transgenic sugar beet plants capable of tolerating herbicide treatment with glyphosate as active ingredient. 
     2. Background of the Related Art 
     Weeds in sugar beet fields are a major problem for the farmer. They compete with the crop thus reducing yield. Today, no single herbicide is able to effectively control all weeds without harming the sugar beet crop itself (Miller et al, J. Sugar Beets Res. 26: 3-4, 1989). In practice, farmers use mixtures of herbicides, which also reduce growth of the crop. Meanwhile the number of weed species having developed resistance to said herbicides continues to increase (Schweizer et al, J. of Sugar Beet Research 28: 1-23, 1991) thereby aggravating the problem of weed control in sugar beet fields. 
     Roundup® is a broadspectrum, environmentally preferable herbicide inhibiting the growth of both weed and crop species. In the context of the present invention one liter of a herbicidal Roundup® solution comprises 360 g of its active ingredient (a.i.) glyphosate (the common name of N-phosponomethyl-glycine) which is taken up by foliage. So far no glyphosate resistant weed has developed in over 20 years of use (Holt et al., Annu. Rev. Plant Physiol., 1993); additionally no natural tolerance to glyphosate has been found in sugar beet. However, pre-emergence use of Roundup® seems to be more efficient for weed control in sugar beet fields than a combination of herbicides often used in sugar beet agriculture, consisting of phenmediphan, metamitron and ethofumesate (Madsen et al, Weed Res. 35: 105-111, 1995). 
     Glyphosate inhibits the biosynthesis of aromatic amino acids, through irreversible binding to 5-enolpyruvylshikimate-3-phosphate synthase (epsps). Within the chloroplast this enzyme catalyzes the reaction of shikimate-3-phosphate and phosphoenolpyruvate to form 5-enolpyruvylshikimate-3-phosphate and phosphate. Approximately one week after application of the herbicide, visible effects can be seen including wilting, yellowing followed by complete browning, deterioration of plant tissue, and decomposition of the roots. 
     To impart glyphosate tolerance to crop species, focus has been on the introduction into plants of epsps genes capable of increasing glyphosate tolerance. Besides plants bacteria and fungi naturally express epsps enzyme activity. The cp4/epsps from Agrobacterium sp. CP4 was found to confer tolerance to glyphosate (Barry et al., “Inhibitors of Amino Acid Biosynthesis: Strategies for Imparting Glyphosate Tolerance to Crop Plants”, in: Biosynthesis and Molecular Regulation of Amino Acids in Plants, Singh et al (eds), American Society of Plant Physiologists, pages 139-145, 1992). Introduction of the cp4/epsps gene into soybean and oilseed rape yielded tolerance to foliar application of the herbicide under field conditions (Delannay et al., Crop Sci. 35: 1461-1467, 1995; Padgette et al., Crop Sci. 35: 1451-1461, 1995). 
     Glyphosate oxidase reductase (gox) isolated from Achromobacter sp. strain LBAA (Barry et al., supra) degrades glyphosate into aminomethyl phosphonic acid, a compound non-toxic for the plant. A combination of the cp4/epsps and glyphosate oxidase (gox) genes has been successfully used to obtain transgenic wheat (Zhou et al., Plant Cell Rep. 15: 159-163, 1995) tolerant to glyphosate. 
     SUMMARY OF THE INVENTION 
     The object of the present invention is to provide sugar beet plants which tolerate glyphosate in doses sufficiently high to effect optimal herbicidal activity. Such plants can be further improved by backcrossing with elite sugar beet lines to optimize agronomic properties such as yield, pathogen resistance, etc. 
     Sugar beets may be transformed using  Agrobacterium tumefaciens  mediated transformation (Fry et al, Third international congress of plant mol. biol., Tuscon, Ariz., USA; D&#39;Halluin et al, Bio/Technology 10: 309-314, 1992; Konwar, J. Plant Biochem &amp; Biotech 3: 37-41,1994). Agrobacterium-mediated transformation often results in more than one copy of the T-DNA being integrated into the plant&#39;s genome. The gene to be integrated is preferably introduced into the T-DNA such that it becomes located close to the T-DNA right border which, contrary to the left border, will almost always be transferred to the plant. 
     Plants according to the present invention tolerate treatment with more than about 3×6 liters of the herbicide Roundup® per hectar (about 18 liters per hectar). The total standard dose to obtain good weed control varies between 4 and 6 liters per hectare, depending on weed pressure. At these concentrations herbicide treatment exerts no detectable effect on plant vigour and leaf chlorosis. The tolerance exhibited by the plants according to the invention is conferred by a transgenically expressed cp4/epsps enzyme activity. A preferred embodiment of the present invention has been deposited with the National Collections of Industrial and Marine Bacteria Limited (NCIMB), 23 St Machar Drive, Aberdeen AB2 1RY, Scotland UK, on Oct. 24, 1997, under the Accession No. 40905. This deposit will be maintained in the NCIMB depository, which is a public depository, for a period of 30 years, or 5 years after the most recent request or for the effective life of the patent, whichever is longer, and will be replaced if it becomes nonviable during that period. Additionally, Applicant has satisfied all the requirements of 37 C.F.R §§1.801-1.809, including providing an indication of the viability of the sample. Applicant imposes no restriction on the availability of the deposited material from the NCIMB; however, Applicant has no authority to waive any restriction imposed by law on the transfer of biological material or its transportation in commerce. 
     The present invention thus relates to a sugar beet plant including the descendants thereof expressing cp4/epsps enzyme activity. In particular the invention relates to a sugar beet plant including the descendants thereof tolerating the treatment with about 4 to about 18 liters Roundup® per hectar. 
     Plants according to the present invention can be obtained by routine Agrobacterium mediated transformation using a transformation vector comprising between right and left T-DNA border sequences a piece of DNA as described in SEQ ID NO: 5 encoding i.a. cp4/epsps. 
     It was surprisingly found within the scope of the present invention, that a transformation event (RRMax) lacking left and right T-DNA border sequences within the transgenic genome and resulting in deletion of a considerable part of the transformation vector DNA while retaining the cp4/epsps encoding DNA provides superior glyphosate tolerance. In particular a piece of DNA as characterized by SEQ ID NO: 1 is found integrated into a highly repetitive region of the genome simlutaneously replacing part of said repetitive genomic sequence. The genomic DNA directly adjacent to that part of the transgene sequence which in the transformation vector used is linked to the T-DNA right border sequence, has the sequence given in SEQ ID NO: 2. The genomic DNA directly adjacent to the other end of the integrated transgenic DNA has the sequence given in SEQ ID NO: 3. The complete DNA sequence of the newly formed genomic DNA arrangement is given in SEQ ID NO: 4. 
     Accordingly, the present invention relates to a sugar beet plant including the descendents thereof wherein DNA characterized by the nucleotide sequence of SEQ ID NO: 1 forms part of the plant&#39;s genome and said nucleotide sequence preferably replaces highly repetitive DNA sequences within the plant&#39;s genome. 
     Preferred herein is a sugar beet plant including the descendents thereof wherein those parts of the genome directly linked to said nulceotide sequence are characterized by the nucleotide sequences of SEQ ID NO: 2 and SEQ ID NO: 3, respectively. 
     The herbicide tolerance engineered into the transgenic seeds and plants mentioned above are passed on by sexual reproduction or vegetative growth and can thus be maintained and propagated in descendant plants. Generally said maintenance and propagation make use of known agricultural methods developed to fit specific purposes such as tilling, sowing or harvesting. As the growing crop is vulnerable to attack and damages caused by insects or infections, measures are undertaken to control plant diseases, insects, nematodes, and other adverse conditions to improve yield. These include mechanical measures such a tillage of the soil or removal of infected plants, as well as the application of agrochemicals such as fungicides, gametocides, nematicides, growth regulants, ripening agents and insecticides. 
     Use of the herbicide tolerance of the transgenic plants and seeds according to the invention can further be made in plant breeding which aims at the development of plants with improved properties such as tolerance of pests, herbicides, or stress, improved nutritional value, increased yield, or improved structure causing less loss from lodging or shattering. The various breeding steps are characterized by well-defined human intervention such as selecting the lines to be crossed, directing pollination of the parental lines, or selecting appropriate descendant plants. Depending on the desired properties different breeding measures are taken. The relevant techniques are well known in the art and include but are not limited to hybridization, inbreeding, backcross breeding, multiline breeding, variety blend, interspecific hybridization, aneuploid techniques, etc. Thus, the transgenic seeds and plants according to the invention can be used for the breeding of improved plant lines which for example increase the effectiveness of conventional methods such as herbicide or pestidice treatment or allow to dispense with said methods due to their modified genetic properties. Alternatively new crops with improved stress tolerance can be obtained which, due to their optimized genetic “equipment”, yield harvested product of better quality than products which were not able to tolerate comparable adverse developmental conditions. 
     In seeds production germination quality and uniformity of seeds are essential product characteristics, whereas germination quality and uniformity of seeds harvested and sold by the farmer is not important. As it is difficult to keep a crop free from other crop and weed seeds, to control seedborne diseases, and to produce seed with good germination, fairly extensive and well-defined seed production practices have been developed by seed producers, who are experienced in the art of growing, conditioning and marketing of pure seed. Thus, it is common practice for the farmer to buy certified seed meeting specific quality standards instead of using seed harvested from his own crop. Propagation material to be used as seeds is customarily treated with a protectant coating comprising herbicides, insecticides, fungicides, bactericides, nematicides, molluscicides or mixtures thereof. Customarily used protectant coatings comprise compounds such as captan, carboxin, thiram (TMTD®), methalaxyl (Apron®), and pirimiphos-methyl (Actellic®). If desired these compounds are formulated together with further carriers, surfactants or application-promoting adjuvants customarily employed in the art of formulation to provide protection against damage caused by bacterial, fungal or animal pests. The protectant coatings may be applied by impregnating propagation material with a liquid formulation or by coating with a combined wet or dry formulation. Other methods of application are also possible such as treatment directed at the buds or the fruit. 
     It is a further aspect of the present invention to provide new agricultural methods such as the methods exemplified above which are characterized by the use of transgenic plants, transgenic plant material, or transgenic seed according to the present invention. 
     In another embodiment, the present invention relates to a transgenic plant cell, tissue, organ, seed or plant part obtained from the transgenic plant. Also included within the invention are transgenic descendants of the plant as well as transgenic plant cells, tissues, organs, seeds and plant parts obtained from the descendants. 
     The invention further relates to a commercial bag containing Roundup® tolerant sugar beet seed capable of expressing cp4/epsps, together with lable instructions for the use thereof. Preferred within this invention is a commercial bag comprising seed of a transgenic plant comprising stably integrated into its genome a piece of DNA having the nucleotide sequence depicted in SEQ ID NO: 1. 
     The transformation methodology employed can be summarized as follows: 
     (a) transforming in vitro grown sugar beet cotyledons using  Agrobacterium tumefaciens  with a vector comprising a piece of DNA encoding cp4/epsps such as described in SEQ ID NO: 5; 
     (b) regenerating shoots in the presence of glyphosate; 
     (c) transferring the shoots to soil in the greenhouse; 
     (d) treating plantlets with glyphosate; 
     (e) visually grading plant vigour and leaf chlorosis; 
     (g) selecting completely normal plants with normal vigour and leafes uneffected by glyphosate treatment; and 
     (h) propagating the selected plants using conventional breeding techniques. 
     In particular shoots are regenerated in the presence of about 0.1 to about 10 mM, preferably about 1 mM glyphosate after 8-12 weeks of selection. Each transgenic shoot is further propagated into ten copies which can optionally be analyzed for the presence of the cp4/epsps transgene using polymerase chain reaction (PCR) before it is transferred to the greenhous for rooting. Light conditions in the greenhouse are 16 hours of light and 8 hours of darkness with a temperature of 22±2° C. At the three to four leaf stage, the plantlets are sprayed with an aqueous solution of Roundup® at a dose of 0.1 to 20 liters, preferably 1 liter per hectar. Visual injury ratings for plant vigour and plant chlorosis based on a scale from 0 (dead plant) to 9 (completely uneffected plant) are taken on individual plants 2 weeks after glyphosate application. Ratings of 0 to 3 are characteristic of susceptible plants. Ratings of 3 to 7 indicate a low to intermediate level of tolerance, and ratings of 8 or 9 indicate good levels of tolerance. In particular the the ratings have the following meaning: 
     
       
         
               
               
             
           
               
                   
               
             
             
               
                 9 
                 Unaffected plant identical to untreated control 
               
               
                 8 
                 Only very small necrosis on the tips of the leaves with less than 
               
               
                   
                 5% of the leaf area affected and yellow 
               
               
                 7 
                 Very small necrosis on the tips of the leaves which start to curl; 
               
               
                   
                 less than 5% of the leaf area are affected and yellow 
               
               
                 6,5,4 
                 Increasing necrosis and leaf curl; leaves are becoming smaller than 
               
               
                   
                 normal 
               
               
                 3,2 
                 No or very limited leaf growth; all leaves are curled and affected 
               
               
                   
                 by necrosis 
               
               
                 1 
                 No growth of the plant; up to 5% of the plant stay green 
               
               
                 0 
                 Dead plant 
               
               
                   
               
             
          
         
       
     
     To collect data from field trials plants with normal vigour and uneffected by glyphosate treatment (rating 9) are further propagated or bred by convential techniques. 
     Transformation is preferably performed using a Ti vector comprising a piece of DNA with the sequence given in SEQ ID NO: 5 containing the cp4/epsps and gox genes both of which have been reported to confer tolerance to glyphosate in certain plant species, and the reporter gene uidA encoding the β-glucuronidase enzyme. The enhanced 35S promoter (Odell et al., Nature 313: 810-812, 1985) is linked to the uidA gene, the figworth mosaic virus (FMV) promoter (Gouwda et al., J. Cell. Biochem. Suppl. 13D: 301, 1989) to the cp4/epsps and gox genes. Upstream of the cp4/epsps and the gox genes a transit peptide (Gasser et al., J. Biol. Chem. 263:4280-4289, 1988) is inserted to achieve targeting of both of the proteins to the chloroplast. 
     One transformation event which was generated in accordance with the present invention is surprisingly found to be uneffected by doses of up to about 3×6 liters (about 18 liters) of the herbicide Roundup® per hectar. Molecular analysis reveals that 
     there is a single copy of transgenic DNA integrated at a single locus; 
     the integrated DNA encodes cp4/epsps and corresponds to truncated vector DNA; 
     the integrated DNA replaces a piece of genomic DNA and has the sequence shown in SEQ ID NO: 1; 
     the genomic DNA directly adjacent to the integrated DNA is characterized by the sequences of SEQ ID NO: 2 and SEQ ID NO: 3 resulting in the new genomic sequence arrangment of SEQ ID NO: 4; 
     the cp4/epsps and the uidA genes are intact whereas the gox gene is truncated; 
     other vector sequences are not present in the transgenic plants. 
     Now that this information is available plants derived from one of the specific transformation events according to the invention can be easily distinguished from other sugar beet plants by means of PCR. Suitable primer pair combinations allow to specifically identify genomic DNA sequences which are only present in plants directly or indirectly resulting from identical transformation events. Such events are not in any way limited to those obtained by Agrobacterium-mediated transformation but may also result from biolistic transformation experiments. 
     The present invention thus further relates to a sugar beet plant including the descendents thereof characterized in that PCR amplification with its genomic DNA as template results in amplification of 
     a 739 bp DNA fragment when using a pair of oligonucleotide primers characterized by the sequences of SEQ ID NO: 18 and SEQ ID NO: 21; or 
     a 834 bp DNA fragment when using a pair of oligonucleotide primers characterized by the sequences of SEQ ID NO: 20 and SEQ ID NO: 24; or 
     a 1057 bp DNA fragment when using a pair of oligonucleotide primers characterized by the sequences of SEQ ID NO: 17 and SEQ ID NO: 22; or 
     a 1224 bp DNA fragment when using a pair of oligonucleotide primers characterized by the sequences of SEQ ID NO: 19 and SEQ ID NO: 25. 
    
    
     EXAMPLES 
     DETAILED DESCRIPTION OF THE INVENTION 
     The following examples further describe the materials and methods used in carrying out the invention and the subsequent results. They are offered by way of illustration, and their recitation should not be considered as a limitation of the claimed invention. 
     Example 1 
     Sugar Beet Transformation 
     Sugar beet of genotype A1012 is transformed with a vector containing a cp4/epsps and a gox gene capable of confering glyphosate tolerance, the nptII gene, which confers resistance to the antibiotic kanamycin and the reporter gene uidA encoding β-glucuronidase (GUS). The nptII and the uidA gene are functionally linked to the enhanced 35S promoter (Odell et al., Nature 313: 810-812, 1985) whereas the cp4/epsps and gox genes are under control of the the figworth mosaic virus (FMV) promoter (Gouwda et al., J. Cell. Biochem. Suppl. 13D: 301, 1989). Additionally the cp4/epsps and gox gene are linked to a transit peptide (Gasser et al., J. Biol. Chem. 263: 4280-4289, 1988) positioned 5′ to target both proteins to the chloroplast. cp4/epsps and uidA make use uf the E9 3′ terminator sequences whereas gox and nptII use the corresponding NOS 3′ sequence. In vitro grown sugar beet (Beta vulgaris. L) are transformed using  A. tumefaciens.  Plants are regenerated as described by Fry et al. (“Genotype-independent transformation of sugarbeet using  Agrobacterium tumefaciens ”, Third international congress of plant mol. biol., Tuscon, Ariz., USA.,1991) and Konwar (J. Plant Biochem &amp; Biotech 3: 37-41, 1994) using glyphosate at a concentration of 1 mM as selection agent. In order to eliminate  A. tumefaciens  the cotyledons are three times incubated in MS medium containing 500 mg/lit cefotaxime (60 min at 45-50 rpm). Regenerated shoots are analyzed after 8-12 weeks of selection on 1 mM glyphosate, 500 mg/lit cefotaxime including passage to fresh media every third week. Each transgenic shoot is further micropropagated into ten copies, analyzed for the presence of the transgene, and transferred to the greenhouse for rooting. 
     Polymerase Chain Reaction (PCR) is used to verify the presence of the cp4/epsps gene. Twenty mg of plant material is collected in an eppendorf tube from regenerated in vitro shoots. Total DNA is extracted essentially according to Tai et al, Plant Mol. Biol. Rep 8: 297-303, 1990. Minor modifications are adding 500 μp of 100 mM Tris-HCl pH 8.0 containing 50 mM Na-EDTA, 1.25% SDS (W/V), 0.38% Na-Bisulfite (W/V), and 200 μg/ml proteinase K and incubating at 65° C. for two hours. Undissolved leaf material is hooked out and total DNA precipitated and frozen for 2 hours at -20° C. PCR is performed according to the instructions included with the Perkin-Elmer Gene-Amp PCR kit (Perkin-Elmer Corp.) using a modified 10×reaction buffer consisiting of 100 mM Tris-HCl pH 8.3, 500 mM KCl, 30 mM MgCl 2 , 1.0% Nonidet P40 (W/V), 0.4 μM cresol red in 24% sucrose (W/V) and the Taq Start Antibody (Clontech). For amplifying cp4/epsps sequences the following primers are used (Shah et al., Science 233: 478-481, 1986): 
     5′-CAC CGG TCT TTT GGA AGG TGA AG-3′ (SEQ ID NO: 6) and 
     5′-AAC GAG ACC CAT AAC GAG GAA GC-3′ (SEQ ID NO: 7). 
     For amplifying a genomic internal control sequence (surA, surB) the following primers are used: 
     5′-AAA CAG TCC CGT GCA TCC CCA AC-3′ (SEQ ID NO: 8) and 
     5′-GAC GCT CTC CTT GAT TCT GTC CC-3′ (SEQ ID NO: 9). 
     No variation in transformation efficiency is seen between different binary plasmids. Micropropagated shoots are transferred to soil in the greenhouse. Light conditions in the greenhouse are 200 μmol m −2  sec −1 , (Osram Power Star HQI-Z, 400 W), 16 hours of light, 8 hours of darkness, and a temperature of 22±2° C. At the three to four leaf stage plantlets derived from 260 independent transformants are treated with Roundup®. A calibrated sprayer is used to apply an aqueous solution of the herbicide at a dose of 1 lit per ha. (360 g.a.i.l −1 ) ). Visual injury ratings for phytotoxic effects, based on a scale from 0 (complete leaf necrosis) to 9 (no visible effects) are taken on individual plants 2 weeks after glyphosate application. For 75 different transformants phytotoxic effects ranging from score 0-6 are detected. The other 185 (71%) transformants score 7 or more which means they show minor or no visible effects after treatment with the herbicide. Those are further crossed with non-transgenic sugar beets to produce F1 progeny for subsequent evaluation in field trials. 
     Field trials are performed to evaluate the different transformation events under field conditions. The plots consist of three rows, each row 9 meter long, with a distance between the rows of 0.5 meter. One individual transformant is drilled in each plot. After an initial Roundup® application of 1 lit per ha (360 g a.i.l −1 ), at the cotyledon stage, to eliminate non-transgenic segregating plants, the rows are singled by hand, to a final stand of 1 plant per 20 centimeter. Each plot is divided in three parts that are treated independently. One plot is treated with the conventional herbicides metamitron 0.1 kg a.i.ha −1 , phenmediphan 0.2 kg a.i.ha −1  and ethofumesate 0.1 kg a.i.ha −1 . The other plot is treated with 2 times 2 lit per ha (1440 g a.i.l −1 ) of Roundup® and the third plot is treated with 2 times 4 liter (2880 g a.i.l −1 ) of Roundup®. The plants are treated at the two leaf stage and at the four leaf stage. Two weeks after the last application, the plants are scored for phytotoxic effects due to herbicide application. Symptoms ranging from complete susceptibility to complete tolerance are found. Differences between scoring in the greenhouse and scoring in the field trial are observed. This is probably due to environmental differences between the greenhouse and the field trial including absence of UVB light under glass. There were no morphological or physiological changes between plants from plots treated with different doses of Roundup®, compared to treatments with a conventional herbicide mixture. Two transformants show high tolerance to Roundup® after treatment with 2 times 2 lit and 2 times 4 lit per ha. 
     After spraying with Roundup®, tolerance was measured by grading for plant vigour (Tr.vig) and leaf chlorosis (Tr.chl). The various grades have the following meaning: 
     9 . . . Unaffected plant identical to untreated control 
     8 . . . Only very small necrosis on the tips of the leaves with less than 5% of the leaf area affected and yellow 
     7 . . . Very small necrosis on the tips of the leaves which start to curl; less than 5% of the leaf area are affected and yellow 
     6,5,4 . . . Increasing necrosis and leaf curl; leaves are becoming smaller than normal 
     3,2 . . . No or very limited leaf growth; all leaves are curled and affected by necrosis 
     1 . . . No growth of the plant; up to 5% of the plant stay green 
     0 . . . Dead plant 
     22 different transformation events (Entry 1-22) and 1 non-transgenic control (Entry 23) are sprayed with 1+4+4 liters of Roundup®/Ha (Total 9 I/Ha). The results of the visual injury rating are given in the following table. 
     
       
         
               
               
               
               
             
               
               
               
               
             
           
               
                   
               
               
                 Entry 
                 Vigour Rating 
                 Chlorosis Rating 
                   
               
               
                   
               
             
             
               
                   
               
             
          
           
               
                 1 
                 9 
                 9 
                 (RRMax) 
               
               
                 2 
                 6 
                 7 
               
               
                 3 
                 2 
                 3 
               
               
                 4 
                 5 
                 7 
               
               
                 5 
                 9 
                 8 
               
               
                 6 
                 3 
                 3 
               
               
                 7 
                 7 
                 7 
               
               
                 8 
                 7 
                 8 
               
               
                 9 
                 4 
                 6 
               
               
                 10 
                 5 
                 6 
               
               
                 11 
                 5 
                 5 
               
               
                 12 
                 4 
                 5 
               
               
                 13 
                 9 
                 9 
               
               
                 14 
                 3 
                 5 
               
               
                 15 
                 4 
                 4 
               
               
                 16 
                 2 
                 3 
               
               
                 17 
                 1 
                 1 
               
               
                 18 
                 2 
                 2 
               
               
                 19 
                 6 
                 8 
               
               
                 20 
                 7 
                 5 
               
               
                 21 
                 4 
                 5 
               
               
                 22 
                 5 
                 7 
               
               
                 23 
                 0 
                 0 
                 (non-transgenic control) 
               
               
                   
               
             
          
         
       
     
     Example 2 
     Copy Number of the Integrated Transgenic DNA 
     The copy number of the transgenic DNA integrated within the genome of the transformant with the highest glyphosate tolerance (RRMax) is determined by Southern blot analysis of restriction fragments extending beyond the right and left border sequences of the transformation vector used. 
     Genomic DNA from RRMax and a non-transgenic plant with the same genetic is isolated from 250 mg of freeze dried sugar beet leaf material according to Saghai-Maroof et al., Proc. Natl. Acad. Sci. USA 81:8014-8018, 1984. DNA of the transformation vector used serves as the positive control. 15 μg of DNA are digested with 120 units of restriction enzyme for 4 hours at 37° C. in a total volume of 400 μl. The digested DNA is precipitated and redissolved in a total volume of 50 μl. Digested DNA, positive plasmid control, and molecular marker size standard (Lambda digested with EcoRI and HindIII) are separated on a 0.8% agarose gel. DNA is visualized with ethidium bromide and a photograph including a ruler is taken. The DNA is then transferred onto a Hybond N+ membrane and hybridized to the probe as described by Ausubel et al. (eds.) in: “Current protocols in molecular biology”, Green Publishing Associates, Inc. and John Wiley &amp; Sons, Inc., New York, 1987. 
     Probes complementary to basepairs 975-1766 of the cp4/epsps gene sequence (SEQ ID NO: 5) and basepairs 7108-7535 of the gox gene sequence (SEQ ID NO: 5) are prepared by Polymerase Chain Reaction (PCR) wherein the transformation vector used serves as the template DNA for the reaction. The cp4/epsps probe is used to determine the number of inserts flanking the right border region. The gox probe is used to determine the number of inserts flanking the left border region. The PCR products are purified using the Geneclean II® Kit of BIO 101 (La Jolla, Calif.). Labelling of the probe with  32 P is achieved making use of the Megaprime™ DNA labelling system of Amersham International (Little Chalfont, UK). 
     Determination of copy number can be performed by analysis of genomic DNA flanking the right border region. Genomic DNA is digested with the restriction enzyme NcoI which cuts the plasmid derived sequences between the 35S promoter and the uidA gene and within the sugar beet genome. The membrane is probed with the internal PCR fragment from the cp4/epsps gene. After digestion with NcoI and hybridizing with the cp4/epsps probe, a single band of 4.7 kb is detected in RRMax. This demonstrates a single copy of transferred DNA at a single locus. 
     Determination of copy number can also be performed by analysis of genomic DNA flanking the left border region. Genomic DNA is digested with the restriction enzyme HindIII which cuts the plasmid derived sequences between the E9 3′ terminator of the uidA gene and the FMV promoter of the gox gene and within the sugar beet genome. After digestion with HindIII and hybridizing with the gox probe, a single band of approximately 2.0 kb is detected for the transformant RRMax. Since the minimum expected fragment size is ≧4.4 kb, the result indicates a truncation of the plasmid DNA during the transfer into the genomic DNA of the plant. Nevertheless, the result confirms that only a single copy of transferred DNA is integrated at a single locus. 
     Example 3 
     Sequence Analysis of the RRMax Integration Site 
     Genomic RRMax DNA is isolated from 250 mg of freezed dried sugar beet leaf material according to Saghai-Maroof et al. supra. A λFIXII phage library with insert sizes of 9-23 kb in an Xhol cloning site is made (ordered with Stratagene) and probed with cp4/epsps and gox probes to screen for putative clones covering the integration event. A total of 25 phages are detected which hybridize to the cp4/epsps or the gox probe. Two clones are found to hybridize to the cp4/epsps probe and the gox probe, respectively. One of them is purified (Fritsch et al., 1987) and further evaluated by Southern blot analysis and PCR. It contains a 15-16 kb insert of genomic DNA including the transgenic DNA and the flanking sugar beet sequences. Said flanking sequences are amplified by PCR using a primer matching FMV promoter or gox gene sequences in combination with a primer matching a sequence in the λFIXII cloning cassette (detailed sequences are given in Table 1 below). The integrated DNA/sugar beet junction regions are sequenced by the Sanger dideoxy-mediated chain-termination method using Applied Biosystems, Model 373A,Version 2.1.1. The primers used for sequencing are given in Table 2. 
     
       
         
               
             
               
               
             
           
               
                 TABLE 1 
               
             
             
               
                   
               
               
                 Primers used for the amplification of the border regions 
               
             
          
           
               
                 Primer 
                 Sequence 
               
               
                   
               
               
                 #3 
                 5′-CAAGAAGGTTGGTATCGCTG-3′ (SEQ ID NO: 10) 
               
               
                 #7 
                 5′-TCTTTTGTGGTCGTCACTGCGTT-3′ (SEQ ID NO: 11) 
               
               
                 #8 
                 5′-GCGAGCTCTAATACGACTCACTAT-3′ (SEQ ID NO: 12) 
               
               
                 #9 
                 5′-CGCGAGCTCAATTAACCCTCACT-3′ (SEQ ID NO: 13) 
               
               
                   
               
             
          
         
       
     
     
       
         
               
             
               
               
               
             
           
               
                 TABLE 2 
               
             
             
               
                   
               
               
                 Sequencing primers 
               
             
          
           
               
                 Primer 
                 Sequence 
                 Description 
               
               
                   
               
               
                 S1 
                 5′-TCTGTACCCTGACCTTGTTG-3′ 
                 to sequence the 
               
               
                   
                 (SEQ ID NO: 14) 
                 flanking right 
               
               
                   
                   
                 border region 
               
               
                 S3 
                 5′-CGTGGATACCACATCGTGAT-3′ 
                 to sequence the 
               
               
                   
                 (SEQ ID NO: 15) 
                 flanking left 
               
               
                   
                   
                 border region 
               
               
                 S4 
                 5′-ACCTTGGCTTGAAGTACTC-3′ 
                 to sequence the 
               
               
                   
                 (SEQ ID NO: 16) 
                 flanking left 
               
               
                   
                   
                 border region 
               
               
                   
               
             
          
         
       
     
     Sequence analysis revealed that the transgenic DNA integrated in the genome of RRMax has the nucleotide sequence given in SEQ ID NO: 1. The integration start point lies between the right border sequence and the FMV promoter and terminates 897 basepairs downstream of the gox startcodon. Translational stop codons for the truncated gox gene can be found 130 and 235 basepairs downstream of the junction site. A HindIII site is identified 210 basepairs downstream and a transcriptional termination signal (AATAAA) 650 basepair downstream of the gox start codon. SEQ ID NO: 2 describes the DNA sequence of the genomic DNA directly linked to the right border region of the transgenic DNA and SEQ ID NO: 3 describes the DNA sequence of the genomic DNA directly linked to the left border region of the transgenic DNA. 
     Example 4 
     Characterization of the Transgenic DNA Integrated by RRMax 
     To further characterize the right border region, genomic DNA of RRMax and transformation vector DNA are digested with either restriction enzyme BamHI, HindIII, BcII or EcoRI. In southern blot analysis the cut DNA is probed with the cp4/epsps PCR fragment described in Example 2. Independent of the restriction enzyme used, digestion with the different restriction enzymes gives rise to a single fragment in Southern blot analysis. The size of the fragments detected are indicated in Table 3. The data demonstrate that a single copy of the DNA was transferred to the plant, and that the DNA transfer into the sugar beet plant resulted in complete transfer of the cp4/epsps gene. The results are in agreement with the results of the copy number determination in example 2 and the nucleotide sequence analysis of example 3. 
     
       
         
               
             
               
               
               
               
               
             
               
               
               
               
               
             
           
               
                 TABLE 3 
               
             
             
               
                   
               
               
                 Southern blot fragment sizes 
               
             
          
           
               
                   
                   
                   
                   
                 transformation 
               
               
                   
                 Enzyme 
                 RRMax 
                   
                 vector 
               
               
                   
                   
               
             
          
           
               
                   
                 BamHI 
                 &gt;10 
                 kb 
                 9.7 kb 
               
               
                   
                 HindIII 
                 2.4 
                 kb 
                 2.4 kb 
               
               
                   
                 BcII 
                 3.2 
                 kb 
                 2.9 kb 
               
               
                   
                 EcoRI 
                 1.8 
                 kb 
                 1.8 kb 
               
               
                   
                   
               
             
          
         
       
     
     To further characterize the piece of transgenic DNA integrated, genomic DNA from RRMax is digested with either restriction enzyme NcoI, BamHI or HindIII. As a control Transformation vector DNA is digested with the same restriction enzymes. The blot is probed with a PCR amplified DNA fragment spanning basepairs 3796-4837 of the uidA gene in SEQ ID NO: 5. The size of the fragments detected is indicated in Table 4. Independent of restriction enzyme used, digestion with the different restriction enzymes gives rise to a single signal on the autoradiographic film demonstrating that the DNA insert has the same characteristics as the internal DNA of the transformation vector used. 
     
       
         
               
             
               
               
               
               
             
           
               
                 TABLE 4 
               
             
             
               
                   
               
               
                 Southern blot fragment sizes 
               
             
          
           
               
                   
                   
                 transformation 
                   
               
               
                   
                 Enzyme 
                 vector 
                 RRMax 
               
               
                   
                   
               
               
                   
                 NcoI 
                 3.4 kb 
                 3.4 kb 
               
               
                   
                 BamHI 
                 3.2 kb 
                 3.2 kb 
               
               
                   
                 HindIII 
                 3.2 kb 
                 3.2 kb 
               
               
                   
                   
               
             
          
         
       
     
     To further characterize the left border region, genomic DNA of RRMax and transformation vector DNA are digested with either restriction enzyme NcoI, BamHI, HindIII or EcoRI. In southern blot analysis the cut DNA is probed with the gox PCR fragment described in Example 2. The size of the fragments detected is indicated in Table 5. Independent of restriction enzyme used, digestion with the different restriction enzymes gives rise to a single fragment fragment on the autoradiographic film. Digestion with either restriction enzyme NcoI, BamHI or EcoRI is expected to identify an internal fragment of a known size. However, none of these restriction enzymes gives rise to such an internal fragment of the expected size. This indicates that the restriction sites for NcoI, BamHI, and EcoRI are absent in the transgenic plant. The results are in agreement with the results obtained by sequencing where it was found that the gox gene was only partly integrated in the plant. Digestion with restriction enzyme HindIII gives rise to a fragment of approximately 2 kb additionally confirming the sequencing data where a HindIII site is located downstream the gox gene in the genome. The results also show that a single copy of the transgenic DNA has been transferred to the plant. They correlates well to the primary results of the copy number determination in example 2 and the nucleotide sequence analysis in example 3. A single copy is integrated in the plant genome whereas the gox gene is partly deleted. 
     
       
         
               
             
               
               
               
               
               
             
               
               
               
               
               
             
           
               
                 TABLE 5 
               
             
             
               
                   
               
               
                 Southern blot fragment sizes 
               
             
          
           
               
                   
                   
                 transformation 
                   
                   
               
               
                   
                 Enzyme 
                 vector 
                 T9100152 
                   
               
               
                   
                   
               
             
          
           
               
                   
                 NcoI 
                 2.5 kb 
                 3.0 
                 kb 
               
               
                   
                 BamHI 
                 2.9 kb 
                 &gt;10 
                 kb 
               
               
                   
                 HindIII 
                 9.5 kb 
                 2.0 
                 kb 
               
               
                   
                 EcoRI 
                 1.6 kb 
                 3.6 
                 kb 
               
               
                   
                   
               
             
          
         
       
     
     Example 5 
     Absence of Other Vector DNA Sequences 
     To verify the absence of oriV, ori-322, aad and nptII sequences in the transformation event RRMax southern blot analysis can be performed using restriction enzyme/probe combinations covering oriV, ori-322, aad and nptII. 
     Transformation vector DNA is digested with restriction enzyme NspI which cuts the plasmid at 17 sites. For the purpose of the analysis a NspI fragment covering oriV and a fragment covering ori-322 and aad are purified and used for Southern Blot analysis. A PCR amplified fragment with the nucleotide sequence given in SEQ ID NO: 26 is used to probe for nptII sequences. All fragments are purified with the Geneclean II® Kit of BIO 101 (La Jolla, Calif.). Labelling with  32 P is achieved with the Megaprime™ DNA labelling system of Amersham International (Little Chalfont, UK). 
     Genomic DNA of RRMax is digested with the restriction enzyme BamHI which cuts the transformation vector DNA at three sites, positioned at basepairs 2321, 5544 and 8413 of SEQ ID NO: 5. Hybridization of the corresponding southern transfer membrane with the probe covering oriV failed to detect a signal. It has to be concluded that oriV is not present in RRMax. 
     Hybridization of the corresponding southern transfer membrane with the probe covering ori-322 and aad failed to detect a signal. It has to be concluded that ori-322 and aad sequences are not present in RRMax. 
     Hybridization of the corresponding southern transfer membrane with the nptII probe also failed to detect a signal, which demonstrates that an nptII gene is not present in line RRMax. 
     Example 6 
     Stability of the RRMax Line 
     Digestion of genomic DNA with the restriction enzyme Bcll and Southern blot analysis using the cp4/epsps gene as a probe are performed as described in Example 2. Four plants from generations 2 and 3, and five plants from generation 4 are used for this analysis. In addition, 3 non-transgenic plants are used as a negative control. Analysis of the primary transformation event results in a single fragment of 3.2 kb. Southern blot analysis of the progeny generations also results in a single fragment of 3.2 kb. If the introduced DNA is stably inherited from generation to generation, the same fragment of 3.2 kb is expected in all plants. 
     Digestion of genomic DNA with HindIII and Southern blot analysis using an internal fragment of the gox gene as a probe are performed as described in Example 2. Analysis of the primary transformation event results in a single fragment of 2.0 kb. Southern blot analysis of the progeny generations also results in a single fragment of 2.0 kb indicating stable inheritance of the trait. 
     Example 7 
     PCR characterization of RRMax 
     Genomic DNA of line RRMax is prepared as described in example 2. About 0.5 μg of DNA are used as template DNA in a PCR reaction using specific combinations of primers characterized by the sequences given in Table 6. Said specific combinations and the size of the fragment amplified are given in Table 7. Depending on the primer pair combination an annealing temperature between 55° C. and 65° C. is used in each of the 30 to 35 amplification cycles. 
     
       
         
               
             
               
               
               
             
           
               
                 TABLE 6 
               
             
             
               
                   
               
               
                 Primer Sequences 
               
             
          
           
               
                 Primer 
                 Sequence 
                 Description 
               
               
                   
               
               
                 A 
                 5′-TCAACCTACAGCTGATTTGGACC-3′ (SEQ ID NO: 17) 
                 near the right border junction 
               
               
                 B 
                 5′-GGACCGGAACGACAATCTGATC-3′ (SEQ ID NO: 18) 
                 near the right border junction 
               
               
                 C 
                 5′-CTAGGGAAGTCCAAATCAGCCG-3′ (SEQ ID NO: 19) 
                 near the left border junction 
               
               
                 D 
                 5′-TTTGGACCGGAACTTTCCAGAAG-3′ (SEQ ID NO: 20) 
                 near the left border junction 
               
               
                 a 
                 5′-CTAACTTGCGCCATCGGAGAAAC-3′ (SEQ ID NO: 21) 
                 within the cp4/epsps gene 
               
               
                 b 
                 5′-GACTTGTCACCTGGAATACGGAC-3′ (SEQ ID NO: 22) 
                 within the cp4/epsps gene 
               
               
                 c 
                 5′-ATTCTTGAGCTCATCAAGCAGCC-3′ (SEQ ID NO: 23) 
                 within the cp4/epsps gene 
               
               
                 e 
                 5′-AAGGTTGGTATCGCTGGAGCTG-3′ (SEQ ID NO: 24) 
                 within the gox sequences 
               
               
                 f 
                 5′-TCTCCACAATGGCTTCCTCTATG-3′ (SEQ ID NO: 25) 
                 within the gox sequences 
               
               
                   
               
             
          
         
       
     
     
       
         
               
               
               
               
             
               
               
               
               
             
           
               
                   
                   
               
               
                   
                 Combination 
                 Size of amplified fragment 
                   
               
               
                   
                   
               
             
             
               
                   
               
             
          
           
               
                   
                 A + a 
                 757 
                 bp (SEQ ID NO:28) 
               
               
                   
                 A + b 
                 1057 
                 bp (SEQ ID NO:29) 
               
               
                   
                 A + c 
                 2352 
                 bp (SEQ ID NO:30) 
               
               
                   
                 B + a 
                 739 
                 bp (SEQ ID NO:27) 
               
               
                   
                 B + b 
                 1039 
                 bp (SEQ ID NO:31) 
               
               
                   
                 B + c 
                 2334 
                 bp (SEQ ID NO:32) 
               
               
                   
                 C + e 
                 934 
                 bp (SEQ ID NO:33) 
               
               
                   
                 C + f 
                 1224 
                 bp (SEQ ID NO:34) 
               
               
                   
                 D + e 
                 888 
                 bp (SEQ ID NO:35) 
               
               
                   
                 D + f 
                 1178 
                 bp (SEQ ID NO:36) 
               
               
                   
                   
               
             
          
         
       
     
     
       
         
           
             36 
           
           
             1 
             8012 
             DNA 
             Sugar beet 
           
            1
aacgacaatc tgatccccat caagcttgag ctcaggattt agcagcattc cagattgggt     60
tcaatcaaca aggtacgagc catatcactt tattcaaatt ggtatcgcca aaaccaagaa    120
ggaactccca tcctcaaagg tttgtaagga agaattctca gtccaaagcc tcaacaaggt    180
cagggtacag agtctccaaa ccattagcca aaagctacag gagatcaatg aagaatcttc    240
aatcaaagta aactactgtt ccagcacatg catcatggtc agtaagtttc agaaaaagac    300
atccaccgaa gacttaaagt tagtgggcat ctttgaaagt aatcttgtca acatcgagca    360
gctggcttgt ggggaccaga caaaaaagga atggtgcaga attgttaggc gcacctacca    420
aaagcatctt tgcctttatt gcaaagataa agcagattcc tctagtacaa gtggggaaca    480
aaataacgtg gaaaagagct gtcctgacag cccactcact aatgcgtatg acgaacgcag    540
tgacgaccac aaaagaattc cctctatata agaaggcatt cattcccatt tgaaggatca    600
tcagatactg aaccaatcct tctagaagat ctaagcttat cgataagctt gatgtaattg    660
gaggaagatc aaaattttca atccccattc ttcgattgct tcaattgaag tttctccgat    720
ggcgcaagtt agcagaatct gcaatggtgt gcagaaccca tctcttatct ccaatctctc    780
gaaatccagt caacgcaaat ctcccttatc ggtttctctg aagacgcagc agcatccacg    840
agcttatccg atttcgtcgt cgtggggatt gaagaagagt gggatgacgt taattggctc    900
tgagcttcgt cctcttaagg tcatgtcttc tgtttccacg gcgtgcatgc ttcacggtgc    960
aagcagccgt ccagcaactg ctcgtaagtc ctctggtctt tctggaaccg tccgtattcc   1020
aggtgacaag tctatctccc acaggtcctt catgtttgga ggtctcgcta gcggtgaaac   1080
tcgtatcacc ggtcttttgg aaggtgaaga tgttatcaac actggtaagg ctatgcaagc   1140
tatgggtgcc agaatccgta aggaaggtga tacttggatc attgatggtg ttggtaacgg   1200
tggactcctt gctcctgagg ctcctctcga tttcggtaac gctgcaactg gttgccgttt   1260
gactatgggt cttgttggtg tttacgattt cgatagcact ttcattggtg acgcttctct   1320
cactaagcgt ccaatgggtc gtgtgttgaa cccacttcgc gaaatgggtg tgcaggtgaa   1380
gtctgaagac ggtgatcgtc ttccagttac cttgcgtgga ccaaagactc caacgccaat   1440
cacctacagg gtacctatgg cttccgctca agtgaagtcc gctgttctgc ttgctggtct   1500
caacacccca ggtatcacca ctgttatcga gccaatcatg actcgtgacc acactgaaaa   1560
gatgcttcaa ggttttggtg ctaaccttac cgttgagact gatgctgacg gtgtgcgtac   1620
catccgtctt gaaggtcgtg gtaagctcac cggtcaagtg attgatgttc caggtgatcc   1680
atcctctact gctttcccat tggttgctgc cttgcttgtt ccaggttccg acgtcaccat   1740
ccttaacgtt ttgatgaacc caacccgtac tggtctcatc ttgactctgc aggaaatggg   1800
tgccgacatc gaagtgatca acccacgtct tgctggtgga gaagacgtgg ctgacttgcg   1860
tgttcgttct tctactttga agggtgttac tgttccagaa gaccgtgctc cttctatgat   1920
cgacgagtat ccaattctcg ctgttgcagc tgcattcgct gaaggtgcta ccgttatgaa   1980
cggtttggaa gaactccgtg ttaaggaaag cgaccgtctt tctgctgtcg caaacggtct   2040
caagctcaac ggtgttgatt gcgatgaagg tgagacttct ctcgtcgtgc gtggtcgtcc   2100
tgacggtaag ggtctcggta acgcttctgg agcagctgtc gctacccacc tcgatcaccg   2160
tatcgctatg agcttcctcg ttatgggtct cgtttctgaa aaccctgtta ctgttgatga   2220
tgctactatg atcgctacta gcttcccaga gttcatggat ttgatggctg gtcttggagc   2280
taagatcgaa ctctccgaca ctaaggctgc ttgatgagct caagaattcg agctcggtac   2340
cggatcctct agctagagct ttcgttcgta tcatcggttt cgacaacgtt cgtcaagttc   2400
aatgcatcag tttcattgcg cacacaccag aatcctactg agttcgagta ttatggcatt   2460
gggaaaactg tttttcttgt accatttgtt gtgcttgtaa tttactgtgt tttttattcg   2520
gttttcgcta tcgaactgtg aaatggaaat ggatggagaa gagttaatga atgatatggt   2580
ccttttgttc attctcaaat taatattatt tgttttttct cttatttgtt gtgtgttgaa   2640
tttgaaatta taagagatat gcaaacattt tgttttgagt aaaaatgtgt caaatcgtgg   2700
cctctaatga ccgaagttaa tatgaggagt aaaacacttg tagttgtacc attatgctta   2760
ttcactaggc aacaaatata ttttcagacc tagaaaagct gcaaatgtta ctgaatacaa   2820
gtatgtcctc ttgtgtttta gacatttatg aactttcctt tatgtaattt tccagaatcc   2880
ttgtcagatt ctaatcattg ctttataatt atagttatac tcatggattt gtagttgagt   2940
atgaaaatat tttttaatgc attttatgac ttgccaattg attgacaaca tgcatcaatc   3000
gacctgcagc cactcgaagc ggccgcgttc aagcttctgc aggtccgatg tgagactttt   3060
caacaaaggg taatatccgg aaacctcctc ggattccatt gcccagctat ctgtcacttt   3120
attgtgaaga tagtggaaaa ggaaggtggc tcctacaaat gccatcattg cgataaagga   3180
aaggccatcg ttgaagatgc ctctgccgac agtggtccca aagatggacc cccacccacg   3240
aggagcatcg tggaaaaaga agacgttcca accacgtctt caaagcaagt ggattgatgt   3300
gatggtccga tgtgagactt ttcaacaaag ggtaatatcc ggaaacctcc tcggattcca   3360
ttgcccagct atctgtcact ttattgtgaa gatagtggaa aaggaaggtg gctcctacaa   3420
atgccatcat tgcgataaag gaaaggccat cgttgaagat gcctctgccg acagtggtcc   3480
caaagatgga cccccaccca cgaggagcat cgtggaaaaa gaagacgttc caaccacgtc   3540
ttcaaagcaa gtggattgat gtgatatctc cactgacgta agggatgacg cacaatccca   3600
ctatccttcg caagaccctt cctctatata aggaagttca tttcatttgg agaggacacg   3660
ctgacaagct gactctagca gatctccatg gtccgtcctg tagaaacccc aacccgtgaa   3720
atcaaaaaac tcgacggcct gtgggcattc agtctggatc gcgaaaactg tggaattgat   3780
cagcgttggt gggaaagcgc gttacaagaa agccgggcaa ttgctgtgcc aggcagtttt   3840
aacgatcagt tcgccgatgc agatattcgt aattatgcgg gcaacgtctg gtatcagcgc   3900
gaagtcttta taccgaaagg ttgggcaggc cagcgtatcg tgctgcgttt cgatgcggtc   3960
actcattacg gcaaagtgtg ggtcaataat caggaagtga tggagcatca gggcggctat   4020
acgccatttg aagccgatgt cacgccgtat gttattgccg ggaaaagtgt acgtatcacc   4080
gtttgtgtga acaacgaact gaactggcag actatcccgc cgggaatggt gattaccgac   4140
gaaaacggca agaaaaagca gtcttacttc catgatttct ttaactatgc cggaatccat   4200
cgcagcgtaa tgctctacac cacgccgaac acctgggtgg acgatatcac cgtggtgacg   4260
catgtcgcgc aagactgtaa ccacgcgtct gttgactggc aggtggtggc caatggtgat   4320
gtcagcgttg aactgcgtga tgcggatcaa caggtggttg caactggaca aggcactagc   4380
gggactttgc aagtggtgaa tccgcacctc tggcaaccgg gtgaaggtta tctctatgaa   4440
ctgtgcgtca cagccaaaag ccagacagag tgtgatatct acccgcttcg cgtcggcatc   4500
cggtcagtgg cagtgaaggg cgaacagttc ctgattaacc acaaaccgtt ctactttact   4560
ggctttggtc gtcatgaaga tgcggactta cgtggcaaag gattcgataa cgtgctgatg   4620
gtgcacgacc acgcattaat ggactggatt ggggccaact cctaccgtac ctcgcattac   4680
ccttacgctg aagagatgct cgactgggca gatgaacatg gcatcgtggt gattgatgaa   4740
actgctgctg tcggctttaa cctctcttta ggcattggtt tcgaagcggg caacaagccg   4800
aaagaactgt acagcgaaga ggcagtcaac ggggaaactc agcaagcgca cttacaggcg   4860
attaaagagc tgatagcgcg tgacaaaaac cacccaagcg tggtgatgtg gagtattgcc   4920
aacgaaccgg atacccgtcc tgcacgggaa tatttcggca tttcgccact ggcggaagca   4980
acgcgtaaac tcgacccgac gcgtccgatc acctgcgtca atgtaatgtt ctgcgacgct   5040
cacaccgata ccatcagcga tctctttgat gtgctgtgcc tgaaccgtta ttacggatgg   5100
tatgtccaaa gcggcgattt ggaaacggca gagaaggtac tggaaaaaga acttctggcc   5160
tggcaggaga aactgcatca gccgattatc atcaccgaat acggcgtgga tacgttagcc   5220
gggctgcact caatgtacac cgacatgtgg agtgaagagt atcagtgtgc atggctggat   5280
atgtatcacc gcgtctttga tcgcgtcagc gccgtcgtcg gtgaacaggt atggaatttc   5340
gccgattttg cgacctcgca aggcatattg cgcgttggcg gtaacaagaa agggatcttc   5400
actcgcgacc gcaaaccgaa gtcggcggct tttctgctgc aaaaacgctg gactggcatg   5460
aacttcggtg aaaaaccgca gcagggaggc aaacaatgaa tcaacaactc tcctggcgca   5520
ccatcgtcgg ctacagcctc ggtggggaat tcgagctcgc ccggggatcc tctagctaga   5580
gctttcgttc gtatcatcgg tttcgacaac gttcgtcaag ttcaatgcat cagtttcatt   5640
gcgcacacac cagaatccta ctgagttcga gtattatggc attgggaaaa ctgtttttct   5700
tgtaccattt gttgtgcttg taatttactg tgttttttat tcggttttcg ctatcgaact   5760
gtgaaatgga aatggatgga gaagagttaa tgaatgatat ggtccttttg ttcattctca   5820
aattaatatt atttgttttt tctcttattt gttgtgtgtt gaatttgaaa ttataagaga   5880
tatgcaaaca ttttgttttg agtaaaaatg tgtcaaatcg tggcctctaa tgaccgaagt   5940
taatatgagg agtaaaacac ttgtagttgt accattatgc ttattcacta ggcaacaaat   6000
atattttcag acctagaaaa gctgcaaatg ttactgaata caagtatgtc ctcttgtgtt   6060
ttagacattt atgaactttc ctttatgtaa ttttccagaa tccttgtcag attctaatca   6120
ttgctttata attatagtta tactcatgga tttgtagttg agtatgaaaa tattttttaa   6180
tgcattttat gacttgccaa ttgattgaca acatgcatca atcgacctgc agcccaagct   6240
tgagctcagg atttagcagc attccagatt gggttcaatc aacaaggtac gagccatatc   6300
actttattca aattggtatc gccaaaacca agaaggaact cccatcctca aaggtttgta   6360
aggaagaatt ctcagtccaa agcctcaaca aggtcagggt acagagtctc caaaccatta   6420
gccaaaagct acaggagatc aatgaagaat cttcaatcaa agtaaactac tgttccagca   6480
catgcatcat ggtcagtaag tttcagaaaa agacatccac cgaagactta aagttagtgg   6540
gcatctttga aagtaatctt gtcaacatcg agcagctggc ttgtggggac cagacaaaaa   6600
aggaatggtg cagaattgtt aggcgcacct accaaaagca tctttgcctt tattgcaaag   6660
ataaagcaga ttcctctagt acaagtgggg aacaaaataa cgtggaaaag agctgtcctg   6720
acagcccact cactaatgcg tatgacgaac gcagtgacga ccacaaaaga attccctcta   6780
tataagaagg cattcattcc catttgaagg atcatcagat actgaaccaa tccttctaga   6840
agatctccac aatggcttcc tctatgctct cttccgctac tatggttgcc tctccggctc   6900
aggccactat ggtcgctcct ttcaacggac ttaagtcctc cgctgccttc ccagccaccc   6960
gcaaggctaa caacgacatt acttccatca caagcaacgg cggaagagtt aactgcatgc   7020
aggtgtggcc tccgattgga aagaagaagt ttgagactct ctcttacctt cctgacctta   7080
ccgattccgg tggtcgcgtc aactgcatgc aggccatggc tgagaaccac aagaaggttg   7140
gtatcgctgg agctggaatc gttggtgttt gcactgcttt gatgcttcaa cgtcgtggat   7200
tcaaggttac cttgattgat ccaaacccac caggtgaagg tgcttctttc ggtaacgctg   7260
gttgcttcaa cggttcctcc gttgttccaa tgtccatgcc aggaaacttg actagcgttc   7320
caaagtggct tcttgaccca atgggtccat tgtccatccg tttcagctac tttccaacca   7380
tcatgccttg gttgattcgt ttcttgcttg ctggaagacc aaacaaggtg aaggagcaag   7440
ctaaggcact ccgtaacctc atcaagtcca ctgtgccttt gatcaagtcc ttggctgagg   7500
aggctgatgc tagccacctt atccgtcacg aaggtcacct taccgtgtac cgtggagaag   7560
cagacttcgc caaggaccgt ggaggttggg aacttcgtcg tctcaacggt gttcgtactc   7620
aaatcctcag cgctgatgca ttgcgtgatt tcgatcctaa cttgtctcac gcctttacca   7680
agggaatcct tatcgaagag aacggtcaca ccatcaaccc acaaggtctc gtgactctct   7740
tgtttcgtcg tttcatcgct aacggtggag agttcgtgtc tgctcgtgtt atcggattcg   7800
agactgaagg tcgtgctctc aagggtatca ccaccaccaa cggtgttctt gctgttgatg   7860
cagctgttgt tgcagctggt gcacactcca agtctcttgc taactccctt ggtgatgaca   7920
tcccattgga taccgaacgt ggataccaca tcgtgatcgc caacccagaa gctgctccac   7980
gtattccaac taccgatgct tctggaaagt tc                                 8012
 
           
             2 
             89 
             DNA 
             Sugar beet 
           
            2
ggattgtgtt tgggttttgt ctgtgtgttt aatgtgttta agggatgaat tagaatgctc     60
ttaatcaacc tacagctgat ttggaccgg                                       89
 
           
             3 
             697 
             DNA 
             Sugar beet 
           
            3
cggtccaaat ttgtttacat tgtgtccaaa tttcggctga tttggacttc cctagctatg     60
ccaactaagc taataaaaaa catgaaacaa caattacaaa ctgtcgagca caccttctac    120
aaactagctt agatttctat tggaagttac aaaacagtaa aactaccaat aggatactaa    180
attaaacata ttaaactatt actcctcaaa agcttgtaca atttgcagaa gaaatgatgg    240
ttgcccaaaa gcttcaaagg gaacctgctg ggaagcctgc tgggacgctg gggatgctgg    300
cagcagcata ccttggcttg aagtactctt ctctcattgg ttttgcttcc cttgcccatg    360
tggtcttcat atggcctcat tacttcccaa gggcttcaaa tcagtaggtg gtggcaacca    420
aaagcatcaa aaacatctcc taaaactagc ttatacaacc ggattacatg agcttatact    480
agcttaactc ttaaagcatg attaacataa tgatgtttaa ggtgtcatta agtattacta    540
atcttgctta agtagagatt aacataggat tagcctaatc aagttgctta agtaaggttt    600
tagaataaac cgagctagtt aggcttaagt agagattaac ataggattag cctaatcaag    660
ttgcttaagt aaggttttag aataaaccga gctagtt                             697
 
           
             4 
             8798 
             DNA 
             Sugar beet 
           
            4
ggattgtgtt tgggttttgt ctgtgtgttt aatgtgttta agggatgaat tagaatgctc     60
ttaatcaacc tacagctgat ttggaccgga acgacaatct gatccccatc aagcttgagc    120
tcaggattta gcagcattcc agattgggtt caatcaacaa ggtacgagcc atatcacttt    180
attcaaattg gtatcgccaa aaccaagaag gaactcccat cctcaaaggt ttgtaaggaa    240
gaattctcag tccaaagcct caacaaggtc agggtacaga gtctccaaac cattagccaa    300
aagctacagg agatcaatga agaatcttca atcaaagtaa actactgttc cagcacatgc    360
atcatggtca gtaagtttca gaaaaagaca tccaccgaag acttaaagtt agtgggcatc    420
tttgaaagta atcttgtcaa catcgagcag ctggcttgtg gggaccagac aaaaaaggaa    480
tggtgcagaa ttgttaggcg cacctaccaa aagcatcttt gcctttattg caaagataaa    540
gcagattcct ctagtacaag tggggaacaa aataacgtgg aaaagagctg tcctgacagc    600
ccactcacta atgcgtatga cgaacgcagt gacgaccaca aaagaattcc ctctatataa    660
gaaggcattc attcccattt gaaggatcat cagatactga accaatcctt ctagaagatc    720
taagcttatc gataagcttg atgtaattgg aggaagatca aaattttcaa tccccattct    780
tcgattgctt caattgaagt ttctccgatg gcgcaagtta gcagaatctg caatggtgtg    840
cagaacccat ctcttatctc caatctctcg aaatccagtc aacgcaaatc tcccttatcg    900
gtttctctga agacgcagca gcatccacga gcttatccga tttcgtcgtc gtggggattg    960
aagaagagtg ggatgacgtt aattggctct gagcttcgtc ctcttaaggt catgtcttct   1020
gtttccacgg cgtgcatgct tcacggtgca agcagccgtc cagcaactgc tcgtaagtcc   1080
tctggtcttt ctggaaccgt ccgtattcca ggtgacaagt ctatctccca caggtccttc   1140
atgtttggag gtctcgctag cggtgaaact cgtatcaccg gtcttttgga aggtgaagat   1200
gttatcaaca ctggtaaggc tatgcaagct atgggtgcca gaatccgtaa ggaaggtgat   1260
acttggatca ttgatggtgt tggtaacggt ggactccttg ctcctgaggc tcctctcgat   1320
ttcggtaacg ctgcaactgg ttgccgtttg actatgggtc ttgttggtgt ttacgatttc   1380
gatagcactt tcattggtga cgcttctctc actaagcgtc caatgggtcg tgtgttgaac   1440
ccacttcgcg aaatgggtgt gcaggtgaag tctgaagacg gtgatcgtct tccagttacc   1500
ttgcgtggac caaagactcc aacgccaatc acctacaggg tacctatggc ttccgctcaa   1560
gtgaagtccg ctgttctgct tgctggtctc aacaccccag gtatcaccac tgttatcgag   1620
ccaatcatga ctcgtgacca cactgaaaag atgcttcaag gttttggtgc taaccttacc   1680
gttgagactg atgctgacgg tgtgcgtacc atccgtcttg aaggtcgtgg taagctcacc   1740
ggtcaagtga ttgatgttcc aggtgatcca tcctctactg ctttcccatt ggttgctgcc   1800
ttgcttgttc caggttccga cgtcaccatc cttaacgttt tgatgaaccc aacccgtact   1860
ggtctcatct tgactctgca ggaaatgggt gccgacatcg aagtgatcaa cccacgtctt   1920
gctggtggag aagacgtggc tgacttgcgt gttcgttctt ctactttgaa gggtgttact   1980
gttccagaag accgtgctcc ttctatgatc gacgagtatc caattctcgc tgttgcagct   2040
gcattcgctg aaggtgctac cgttatgaac ggtttggaag aactccgtgt taaggaaagc   2100
gaccgtcttt ctgctgtcgc aaacggtctc aagctcaacg gtgttgattg cgatgaaggt   2160
gagacttctc tcgtcgtgcg tggtcgtcct gacggtaagg gtctcggtaa cgcttctgga   2220
gcagctgtcg ctacccacct cgatcaccgt atcgctatga gcttcctcgt tatgggtctc   2280
gtttctgaaa accctgttac tgttgatgat gctactatga tcgctactag cttcccagag   2340
ttcatggatt tgatggctgg tcttggagct aagatcgaac tctccgacac taaggctgct   2400
tgatgagctc aagaattcga gctcggtacc ggatcctcta gctagagctt tcgttcgtat   2460
catcggtttc gacaacgttc gtcaagttca atgcatcagt ttcattgcgc acacaccaga   2520
atcctactga gttcgagtat tatggcattg ggaaaactgt ttttcttgta ccatttgttg   2580
tgcttgtaat ttactgtgtt ttttattcgg ttttcgctat cgaactgtga aatggaaatg   2640
gatggagaag agttaatgaa tgatatggtc cttttgttca ttctcaaatt aatattattt   2700
gttttttctc ttatttgttg tgtgttgaat ttgaaattat aagagatatg caaacatttt   2760
gttttgagta aaaatgtgtc aaatcgtggc ctctaatgac cgaagttaat atgaggagta   2820
aaacacttgt agttgtacca ttatgcttat tcactaggca acaaatatat tttcagacct   2880
agaaaagctg caaatgttac tgaatacaag tatgtcctct tgtgttttag acatttatga   2940
actttccttt atgtaatttt ccagaatcct tgtcagattc taatcattgc tttataatta   3000
tagttatact catggatttg tagttgagta tgaaaatatt ttttaatgca ttttatgact   3060
tgccaattga ttgacaacat gcatcaatcg acctgcagcc actcgaagcg gccgcgttca   3120
agcttctgca ggtccgatgt gagacttttc aacaaagggt aatatccgga aacctcctcg   3180
gattccattg cccagctatc tgtcacttta ttgtgaagat agtggaaaag gaaggtggct   3240
cctacaaatg ccatcattgc gataaaggaa aggccatcgt tgaagatgcc tctgccgaca   3300
gtggtcccaa agatggaccc ccacccacga ggagcatcgt ggaaaaagaa gacgttccaa   3360
ccacgtcttc aaagcaagtg gattgatgtg atggtccgat gtgagacttt tcaacaaagg   3420
gtaatatccg gaaacctcct cggattccat tgcccagcta tctgtcactt tattgtgaag   3480
atagtggaaa aggaaggtgg ctcctacaaa tgccatcatt gcgataaagg aaaggccatc   3540
gttgaagatg cctctgccga cagtggtccc aaagatggac ccccacccac gaggagcatc   3600
gtggaaaaag aagacgttcc aaccacgtct tcaaagcaag tggattgatg tgatatctcc   3660
actgacgtaa gggatgacgc acaatcccac tatccttcgc aagacccttc ctctatataa   3720
ggaagttcat ttcatttgga gaggacacgc tgacaagctg actctagcag atctccatgg   3780
tccgtcctgt agaaacccca acccgtgaaa tcaaaaaact cgacggcctg tgggcattca   3840
gtctggatcg cgaaaactgt ggaattgatc agcgttggtg ggaaagcgcg ttacaagaaa   3900
gccgggcaat tgctgtgcca ggcagtttta acgatcagtt cgccgatgca gatattcgta   3960
attatgcggg caacgtctgg tatcagcgcg aagtctttat accgaaaggt tgggcaggcc   4020
agcgtatcgt gctgcgtttc gatgcggtca ctcattacgg caaagtgtgg gtcaataatc   4080
aggaagtgat ggagcatcag ggcggctata cgccatttga agccgatgtc acgccgtatg   4140
ttattgccgg gaaaagtgta cgtatcaccg tttgtgtgaa caacgaactg aactggcaga   4200
ctatcccgcc gggaatggtg attaccgacg aaaacggcaa gaaaaagcag tcttacttcc   4260
atgatttctt taactatgcc ggaatccatc gcagcgtaat gctctacacc acgccgaaca   4320
cctgggtgga cgatatcacc gtggtgacgc atgtcgcgca agactgtaac cacgcgtctg   4380
ttgactggca ggtggtggcc aatggtgatg tcagcgttga actgcgtgat gcggatcaac   4440
aggtggttgc aactggacaa ggcactagcg ggactttgca agtggtgaat ccgcacctct   4500
ggcaaccggg tgaaggttat ctctatgaac tgtgcgtcac agccaaaagc cagacagagt   4560
gtgatatcta cccgcttcgc gtcggcatcc ggtcagtggc agtgaagggc gaacagttcc   4620
tgattaacca caaaccgttc tactttactg gctttggtcg tcatgaagat gcggacttac   4680
gtggcaaagg attcgataac gtgctgatgg tgcacgacca cgcattaatg gactggattg   4740
gggccaactc ctaccgtacc tcgcattacc cttacgctga agagatgctc gactgggcag   4800
atgaacatgg catcgtggtg attgatgaaa ctgctgctgt cggctttaac ctctctttag   4860
gcattggttt cgaagcgggc aacaagccga aagaactgta cagcgaagag gcagtcaacg   4920
gggaaactca gcaagcgcac ttacaggcga ttaaagagct gatagcgcgt gacaaaaacc   4980
acccaagcgt ggtgatgtgg agtattgcca acgaaccgga tacccgtcct gcacgggaat   5040
atttcggcat ttcgccactg gcggaagcaa cgcgtaaact cgacccgacg cgtccgatca   5100
cctgcgtcaa tgtaatgttc tgcgacgctc acaccgatac catcagcgat ctctttgatg   5160
tgctgtgcct gaaccgttat tacggatggt atgtccaaag cggcgatttg gaaacggcag   5220
agaaggtact ggaaaaagaa cttctggcct ggcaggagaa actgcatcag ccgattatca   5280
tcaccgaata cggcgtggat acgttagccg ggctgcactc aatgtacacc gacatgtgga   5340
gtgaagagta tcagtgtgca tggctggata tgtatcaccg cgtctttgat cgcgtcagcg   5400
ccgtcgtcgg tgaacaggta tggaatttcg ccgattttgc gacctcgcaa ggcatattgc   5460
gcgttggcgg taacaagaaa gggatcttca ctcgcgaccg caaaccgaag tcggcggctt   5520
ttctgctgca aaaacgctgg actggcatga acttcggtga aaaaccgcag cagggaggca   5580
aacaatgaat caacaactct cctggcgcac catcgtcggc tacagcctcg gtggggaatt   5640
cgagctcgcc cggggatcct ctagctagag ctttcgttcg tatcatcggt ttcgacaacg   5700
ttcgtcaagt tcaatgcatc agtttcattg cgcacacacc agaatcctac tgagttcgag   5760
tattatggca ttgggaaaac tgtttttctt gtaccatttg ttgtgcttgt aatttactgt   5820
gttttttatt cggttttcgc tatcgaactg tgaaatggaa atggatggag aagagttaat   5880
gaatgatatg gtccttttgt tcattctcaa attaatatta tttgtttttt ctcttatttg   5940
ttgtgtgttg aatttgaaat tataagagat atgcaaacat tttgttttga gtaaaaatgt   6000
gtcaaatcgt ggcctctaat gaccgaagtt aatatgagga gtaaaacact tgtagttgta   6060
ccattatgct tattcactag gcaacaaata tattttcaga cctagaaaag ctgcaaatgt   6120
tactgaatac aagtatgtcc tcttgtgttt tagacattta tgaactttcc tttatgtaat   6180
tttccagaat ccttgtcaga ttctaatcat tgctttataa ttatagttat actcatggat   6240
ttgtagttga gtatgaaaat attttttaat gcattttatg acttgccaat tgattgacaa   6300
catgcatcaa tcgacctgca gcccaagctt gagctcagga tttagcagca ttccagattg   6360
ggttcaatca acaaggtacg agccatatca ctttattcaa attggtatcg ccaaaaccaa   6420
gaaggaactc ccatcctcaa aggtttgtaa ggaagaattc tcagtccaaa gcctcaacaa   6480
ggtcagggta cagagtctcc aaaccattag ccaaaagcta caggagatca atgaagaatc   6540
ttcaatcaaa gtaaactact gttccagcac atgcatcatg gtcagtaagt ttcagaaaaa   6600
gacatccacc gaagacttaa agttagtggg catctttgaa agtaatcttg tcaacatcga   6660
gcagctggct tgtggggacc agacaaaaaa ggaatggtgc agaattgtta ggcgcaccta   6720
ccaaaagcat ctttgccttt attgcaaaga taaagcagat tcctctagta caagtgggga   6780
acaaaataac gtggaaaaga gctgtcctga cagcccactc actaatgcgt atgacgaacg   6840
cagtgacgac cacaaaagaa ttccctctat ataagaaggc attcattccc atttgaagga   6900
tcatcagata ctgaaccaat ccttctagaa gatctccaca atggcttcct ctatgctctc   6960
ttccgctact atggttgcct ctccggctca ggccactatg gtcgctcctt tcaacggact   7020
taagtcctcc gctgccttcc cagccacccg caaggctaac aacgacatta cttccatcac   7080
aagcaacggc ggaagagtta actgcatgca ggtgtggcct ccgattggaa agaagaagtt   7140
tgagactctc tcttaccttc ctgaccttac cgattccggt ggtcgcgtca actgcatgca   7200
ggccatggct gagaaccaca agaaggttgg tatcgctgga gctggaatcg ttggtgtttg   7260
cactgctttg atgcttcaac gtcgtggatt caaggttacc ttgattgatc caaacccacc   7320
aggtgaaggt gcttctttcg gtaacgctgg ttgcttcaac ggttcctccg ttgttccaat   7380
gtccatgcca ggaaacttga ctagcgttcc aaagtggctt cttgacccaa tgggtccatt   7440
gtccatccgt ttcagctact ttccaaccat catgccttgg ttgattcgtt tcttgcttgc   7500
tggaagacca aacaaggtga aggagcaagc taaggcactc cgtaacctca tcaagtccac   7560
tgtgcctttg atcaagtcct tggctgagga ggctgatgct agccacctta tccgtcacga   7620
aggtcacctt accgtgtacc gtggagaagc agacttcgcc aaggaccgtg gaggttggga   7680
acttcgtcgt ctcaacggtg ttcgtactca aatcctcagc gctgatgcat tgcgtgattt   7740
cgatcctaac ttgtctcacg cctttaccaa gggaatcctt atcgaagaga acggtcacac   7800
catcaaccca caaggtctcg tgactctctt gtttcgtcgt ttcatcgcta acggtggaga   7860
gttcgtgtct gctcgtgtta tcggattcga gactgaaggt cgtgctctca agggtatcac   7920
caccaccaac ggtgttcttg ctgttgatgc agctgttgtt gcagctggtg cacactccaa   7980
gtctcttgct aactcccttg gtgatgacat cccattggat accgaacgtg gataccacat   8040
cgtgatcgcc aacccagaag ctgctccacg tattccaact accgatgctt ctggaaagtt   8100
ccggtccaaa tttgtttaca ttgtgtccaa atttcggctg atttggactt ccctagctat   8160
gccaactaag ctaataaaaa acatgaaaca acaattacaa actgtcgagc acaccttcta   8220
caaactagct tagatttcta ttggaagtta caaaacagta aaactaccaa taggatacta   8280
aattaaacat attaaactat tactcctcaa aagcttgtac aatttgcaga agaaatgatg   8340
gttgcccaaa agcttcaaag ggaacctgct gggaagcctg ctgggacgct ggggatgctg   8400
gcagcagcat accttggctt gaagtactct tctctcattg gttttgcttc ccttgcccat   8460
gtggtcttca tatggcctca ttacttccca agggcttcaa atcagtaggt ggtggcaacc   8520
aaaagcatca aaaacatctc ctaaaactag cttatacaac cggattacat gagcttatac   8580
tagcttaact cttaaagcat gattaacata atgatgttta aggtgtcatt aagtattact   8640
aatcttgctt aagtagagat taacatagga ttagcctaat caagttgctt aagtaaggtt   8700
ttagaataaa ccgagctagt taggcttaag tagagattaa cataggatta gcctaatcaa   8760
gttgcttaag taaggtttta gaataaaccg agctagtt                           8798
 
           
             5 
             8418 
             DNA 
             Sugar beet 
           
            5
aagcttgagc tcaggattta gcagcattcc agattgggtt caatcaacaa ggtacgagcc     60
atatcacttt attcaaattg gtatcgccaa aaccaagaag gaactcccat cctcaaaggt    120
ttgtaaggaa gaattctcag tccaaagcct caacaaggtc agggtacaga gtctccaaac    180
cattagccaa aagctacagg agatcaatga agaatcttca atcaaagtaa actactgttc    240
cagcacatgc atcatggtca gtaagtttca gaaaaagaca tccaccgaag acttaaagtt    300
agtgggcatc tttgaaagta atcttgtcaa catcgagcag ctggcttgtg gggaccagac    360
aaaaaaggaa tggtgcagaa ttgttaggcg cacctaccaa aagcatcttt gcctttattg    420
caaagataaa gcagattcct ctagtacaag tggggaacaa aataacgtgg aaaagagctg    480
tcctgacagc ccactcacta atgcgtatga cgaacgcagt gacgaccaca aaagaattcc    540
ctctatataa gaaggcattc attcccattt gaaggatcat cagatactga accaatcctt    600
ctagaagatc taagcttatc gataagcttg atgtaattgg aggaagatca aaattttcaa    660
tccccattct tcgattgctt caattgaagt ttctccgatg gcgcaagtta gcagaatctg    720
caatggtgtg cagaacccat ctcttatctc caatctctcg aaatccagtc aacgcaaatc    780
tcccttatcg gtttctctga agacgcagca gcatccacga gcttatccga tttcgtcgtc    840
gtggggattg aagaagagtg ggatgacgtt aattggctct gagcttcgtc ctcttaaggt    900
catgtcttct gtttccacgg cgtgcatgct tcacggtgca agcagccgtc cagcaactgc    960
tcgtaagtcc tctggtcttt ctggaaccgt ccgtattcca ggtgacaagt ctatctccca   1020
caggtccttc atgtttggag gtctcgctag cggtgaaact cgtatcaccg gtcttttgga   1080
aggtgaagat gttatcaaca ctggtaaggc tatgcaagct atgggtgcca gaatccgtaa   1140
ggaaggtgat acttggatca ttgatggtgt tggtaacggt ggactccttg ctcctgaggc   1200
tcctctcgat ttcggtaacg ctgcaactgg ttgccgtttg actatgggtc ttgttggtgt   1260
ttacgatttc gatagcactt tcattggtga cgcttctctc actaagcgtc caatgggtcg   1320
tgtgttgaac ccacttcgcg aaatgggtgt gcaggtgaag tctgaagacg gtgatcgtct   1380
tccagttacc ttgcgtggac caaagactcc aacgccaatc acctacaggg tacctatggc   1440
ttccgctcaa gtgaagtccg ctgttctgct tgctggtctc aacaccccag gtatcaccac   1500
tgttatcgag ccaatcatga ctcgtgacca cactgaaaag atgcttcaag gttttggtgc   1560
taaccttacc gttgagactg atgctgacgg tgtgcgtacc atccgtcttg aaggtcgtgg   1620
taagctcacc ggtcaagtga ttgatgttcc aggtgatcca tcctctactg ctttcccatt   1680
ggttgctgcc ttgcttgttc caggttccga cgtcaccatc cttaacgttt tgatgaaccc   1740
aacccgtact ggtctcatct tgactctgca ggaaatgggt gccgacatcg aagtgatcaa   1800
cccacgtctt gctggtggag aagacgtggc tgacttgcgt gttcgttctt ctactttgaa   1860
gggtgttact gttccagaag accgtgctcc ttctatgatc gacgagtatc caattctcgc   1920
tgttgcagct gcattcgctg aaggtgctac cgttatgaac ggtttggaag aactccgtgt   1980
taaggaaagc gaccgtcttt ctgctgtcgc aaacggtctc aagctcaacg gtgttgattg   2040
cgatgaaggt gagacttctc tcgtcgtgcg tggtcgtcct gacggtaagg gtctcggtaa   2100
cgcttctgga gcagctgtcg ctacccacct cgatcaccgt atcgctatga gcttcctcgt   2160
tatgggtctc gtttctgaaa accctgttac tgttgatgat gctactatga tcgctactag   2220
cttcccagag ttcatggatt tgatggctgg tcttggagct aagatcgaac tctccgacac   2280
taaggctgct tgatgagctc aagaattcga gctcggtacc ggatcctcta gctagagctt   2340
tcgttcgtat catcggtttc gacaacgttc gtcaagttca atgcatcagt ttcattgcgc   2400
acacaccaga atcctactga gttcgagtat tatggcattg ggaaaactgt ttttcttgta   2460
ccatttgttg tgcttgtaat ttactgtgtt ttttattcgg ttttcgctat cgaactgtga   2520
aatggaaatg gatggagaag agttaatgaa tgatatggtc cttttgttca ttctcaaatt   2580
aatattattt gttttttctc ttatttgttg tgtgttgaat ttgaaattat aagagatatg   2640
caaacatttt gttttgagta aaaatgtgtc aaatcgtggc ctctaatgac cgaagttaat   2700
atgaggagta aaacacttgt agttgtacca ttatgcttat tcactaggca acaaatatat   2760
tttcagacct agaaaagctg caaatgttac tgaatacaag tatgtcctct tgtgttttag   2820
acatttatga actttccttt atgtaatttt ccagaatcct tgtcagattc taatcattgc   2880
tttataatta tagttatact catggatttg tagttgagta tgaaaatatt ttttaatgca   2940
ttttatgact tgccaattga ttgacaacat gcatcaatcg acctgcagcc actcgaagcg   3000
gccgcgttca agcttctgca ggtccgatgt gagacttttc aacaaagggt aatatccgga   3060
aacctcctcg gattccattg cccagctatc tgtcacttta ttgtgaagat agtggaaaag   3120
gaaggtggct cctacaaatg ccatcattgc gataaaggaa aggccatcgt tgaagatgcc   3180
tctgccgaca gtggtcccaa agatggaccc ccacccacga ggagcatcgt ggaaaaagaa   3240
gacgttccaa ccacgtcttc aaagcaagtg gattgatgtg atggtccgat gtgagacttt   3300
tcaacaaagg gtaatatccg gaaacctcct cggattccat tgcccagcta tctgtcactt   3360
tattgtgaag atagtggaaa aggaaggtgg ctcctacaaa tgccatcatt gcgataaagg   3420
aaaggccatc gttgaagatg cctctgccga cagtggtccc aaagatggac ccccacccac   3480
gaggagcatc gtggaaaaag aagacgttcc aaccacgtct tcaaagcaag tggattgatg   3540
tgatatctcc actgacgtaa gggatgacgc acaatcccac tatccttcgc aagacccttc   3600
ctctatataa ggaagttcat ttcatttgga gaggacacgc tgacaagctg actctagcag   3660
atctccatgg tccgtcctgt agaaacccca acccgtgaaa tcaaaaaact cgacggcctg   3720
tgggcattca gtctggatcg cgaaaactgt ggaattgatc agcgttggtg ggaaagcgcg   3780
ttacaagaaa gccgggcaat tgctgtgcca ggcagtttta acgatcagtt cgccgatgca   3840
gatattcgta attatgcggg caacgtctgg tatcagcgcg aagtctttat accgaaaggt   3900
tgggcaggcc agcgtatcgt gctgcgtttc gatgcggtca ctcattacgg caaagtgtgg   3960
gtcaataatc aggaagtgat ggagcatcag ggcggctata cgccatttga agccgatgtc   4020
acgccgtatg ttattgccgg gaaaagtgta cgtatcaccg tttgtgtgaa caacgaactg   4080
aactggcaga ctatcccgcc gggaatggtg attaccgacg aaaacggcaa gaaaaagcag   4140
tcttacttcc atgatttctt taactatgcc ggaatccatc gcagcgtaat gctctacacc   4200
acgccgaaca cctgggtgga cgatatcacc gtggtgacgc atgtcgcgca agactgtaac   4260
cacgcgtctg ttgactggca ggtggtggcc aatggtgatg tcagcgttga actgcgtgat   4320
gcggatcaac aggtggttgc aactggacaa ggcactagcg ggactttgca agtggtgaat   4380
ccgcacctct ggcaaccggg tgaaggttat ctctatgaac tgtgcgtcac agccaaaagc   4440
cagacagagt gtgatatcta cccgcttcgc gtcggcatcc ggtcagtggc agtgaagggc   4500
gaacagttcc tgattaacca caaaccgttc tactttactg gctttggtcg tcatgaagat   4560
gcggacttac gtggcaaagg attcgataac gtgctgatgg tgcacgacca cgcattaatg   4620
gactggattg gggccaactc ctaccgtacc tcgcattacc cttacgctga agagatgctc   4680
gactgggcag atgaacatgg catcgtggtg attgatgaaa ctgctgctgt cggctttaac   4740
ctctctttag gcattggttt cgaagcgggc aacaagccga aagaactgta cagcgaagag   4800
gcagtcaacg gggaaactca gcaagcgcac ttacaggcga ttaaagagct gatagcgcgt   4860
gacaaaaacc acccaagcgt ggtgatgtgg agtattgcca acgaaccgga tacccgtcct   4920
gcacgggaat atttcggcat ttcgccactg gcggaagcaa cgcgtaaact cgacccgacg   4980
cgtccgatca cctgcgtcaa tgtaatgttc tgcgacgctc acaccgatac catcagcgat   5040
ctctttgatg tgctgtgcct gaaccgttat tacggatggt atgtccaaag cggcgatttg   5100
gaaacggcag agaaggtact ggaaaaagaa cttctggcct ggcaggagaa actgcatcag   5160
ccgattatca tcaccgaata cggcgtggat acgttagccg ggctgcactc aatgtacacc   5220
gacatgtgga gtgaagagta tcagtgtgca tggctggata tgtatcaccg cgtctttgat   5280
cgcgtcagcg ccgtcgtcgg tgaacaggta tggaatttcg ccgattttgc gacctcgcaa   5340
ggcatattgc gcgttggcgg taacaagaaa gggatcttca ctcgcgaccg caaaccgaag   5400
tcggcggctt ttctgctgca aaaacgctgg actggcatga acttcggtga aaaaccgcag   5460
cagggaggca aacaatgaat caacaactct cctggcgcac catcgtcggc tacagcctcg   5520
gtggggaatt cgagctcgcc cggggatcct ctagctagag ctttcgttcg tatcatcggt   5580
ttcgacaacg ttcgtcaagt tcaatgcatc agtttcattg cgcacacacc agaatcctac   5640
tgagttcgag tattatggca ttgggaaaac tgtttttctt gtaccatttg ttgtgcttgt   5700
aatttactgt gttttttatt cggttttcgc tatcgaactg tgaaatggaa atggatggag   5760
aagagttaat gaatgatatg gtccttttgt tcattctcaa attaatatta tttgtttttt   5820
ctcttatttg ttgtgtgttg aatttgaaat tataagagat atgcaaacat tttgttttga   5880
gtaaaaatgt gtcaaatcgt ggcctctaat gaccgaagtt aatatgagga gtaaaacact   5940
tgtagttgta ccattatgct tattcactag gcaacaaata tattttcaga cctagaaaag   6000
ctgcaaatgt tactgaatac aagtatgtcc tcttgtgttt tagacattta tgaactttcc   6060
tttatgtaat tttccagaat ccttgtcaga ttctaatcat tgctttataa ttatagttat   6120
actcatggat ttgtagttga gtatgaaaat attttttaat gcattttatg acttgccaat   6180
tgattgacaa catgcatcaa tcgacctgca gcccaagctt gagctcagga tttagcagca   6240
ttccagattg ggttcaatca acaaggtacg agccatatca ctttattcaa attggtatcg   6300
ccaaaaccaa gaaggaactc ccatcctcaa aggtttgtaa ggaagaattc tcagtccaaa   6360
gcctcaacaa ggtcagggta cagagtctcc aaaccattag ccaaaagcta caggagatca   6420
atgaagaatc ttcaatcaaa gtaaactact gttccagcac atgcatcatg gtcagtaagt   6480
ttcagaaaaa gacatccacc gaagacttaa agttagtggg catctttgaa agtaatcttg   6540
tcaacatcga gcagctggct tgtggggacc agacaaaaaa ggaatggtgc agaattgtta   6600
ggcgcaccta ccaaaagcat ctttgccttt attgcaaaga taaagcagat tcctctagta   6660
caagtgggga acaaaataac gtggaaaaga gctgtcctga cagcccactc actaatgcgt   6720
atgacgaacg cagtgacgac cacaaaagaa ttccctctat ataagaaggc attcattccc   6780
atttgaagga tcatcagata ctgaaccaat ccttctagaa gatctccaca atggcttcct   6840
ctatgctctc ttccgctact atggttgcct ctccggctca ggccactatg gtcgctcctt   6900
tcaacggact taagtcctcc gctgccttcc cagccacccg caaggctaac aacgacatta   6960
cttccatcac aagcaacggc ggaagagtta actgcatgca ggtgtggcct ccgattggaa   7020
agaagaagtt tgagactctc tcttaccttc ctgaccttac cgattccggt ggtcgcgtca   7080
actgcatgca ggccatggct gagaaccaca agaaggttgg tatcgctgga gctggaatcg   7140
ttggtgtttg cactgctttg atgcttcaac gtcgtggatt caaggttacc ttgattgatc   7200
caaacccacc aggtgaaggt gcttctttcg gtaacgctgg ttgcttcaac ggttcctccg   7260
ttgttccaat gtccatgcca ggaaacttga ctagcgttcc aaagtggctt cttgacccaa   7320
tgggtccatt gtccatccgt ttcagctact ttccaaccat catgccttgg ttgattcgtt   7380
tcttgcttgc tggaagacca aacaaggtga aggagcaagc taaggcactc cgtaacctca   7440
tcaagtccac tgtgcctttg atcaagtcct tggctgagga ggctgatgct agccacctta   7500
tccgtcacga aggtcacctt accgtgtacc gtggagaagc agacttcgcc aaggaccgtg   7560
gaggttggga acttcgtcgt ctcaacggtg ttcgtactca aatcctcagc gctgatgcat   7620
tgcgtgattt cgatcctaac ttgtctcacg cctttaccaa gggaatcctt atcgaagaga   7680
acggtcacac catcaaccca caaggtctcg tgactctctt gtttcgtcgt ttcatcgcta   7740
acggtggaga gttcgtgtct gctcgtgtta tcggattcga gactgaaggt cgtgctctca   7800
agggtatcac caccaccaac ggtgttcttg ctgttgatgc agctgttgtt gcagctggtg   7860
cacactccaa gtctcttgct aactcccttg gtgatgacat cccattggat accgaacgtg   7920
gataccacat cgtgatcgcc aacccagaag ctgctccacg tattccaact accgatgctt   7980
ctggaaagtt catcgctact cctatggaga tgggtcttcg tgttgctgga accgttgagt   8040
tcgctggtct cactgctgct cctaactgga agcgtgctca cgttctctac actcacgctc   8100
gtaagttgct tccagctctc gctcctgcca gttctgaaga acgttactcc aagtggatgg   8160
gtttccgtcc aagcatccca gattcccttc cagtgattgg tcgtgctacc cgtactccag   8220
acgttatcta cgctttcggt cacggtcacc tcggtatgac tggtgctcca atgaccgcaa   8280
ccctcgtttc tgagctcctc gcaggtgaga agacctctat cgacatctct ccattcgcac   8340
caaaccgttt cggtattggt aagtccaagc aaactggtcc tgcatcctaa gtgggaattc   8400
gagctcggta ccggatcc                                                 8418
 
           
             6 
             23 
             DNA 
             Artificial sequence 
             
               misc_feature 
               (1)..(23) 
               Primer 
             
           
            6
caccggtctt ttggaaggtg aag                                             23
 
           
             7 
             23 
             DNA 
             Artificial sequence 
             
               misc_feature 
               (1)..(23) 
               Primer 
             
           
            7
aacgagaccc ataacgagga agc                                             23
 
           
             8 
             23 
             DNA 
             Articifial sequence 
             
               misc_feature 
               (1)..(23) 
               Primer 
             
           
            8
aaacagtccc gtgcatcccc aac                                             23
 
           
             9 
             23 
             DNA 
             Artificial sequence 
             
               misc_feature 
               (1)..(23) 
               Pimer 
             
           
            9
gacgctctcc ttgattctgt ccc                                             23
 
           
             10 
             20 
             DNA 
             Artificial sequence 
             
               misc_feature 
               (1)..(20) 
               Primer 
             
           
            10
caagaaggtt ggtatcgctg                                                 20
 
           
             11 
             23 
             DNA 
             Artificial sequence 
             
               misc_feature 
               (1)..(23) 
               Primer 
             
           
            11
tcttttgtgg tcgtcactgc gtt                                             23
 
           
             12 
             24 
             DNA 
             Artificial sequence 
             
               misc_feature 
               (1)..(24) 
               Primer 
             
           
            12
gcgagctcta atacgactca ctat                                            24
 
           
             13 
             23 
             DNA 
             Artificial sequence 
             
               misc_feature 
               (1)..(23) 
               Primer 
             
           
            13
cgcgagctca attaaccctc act                                             23
 
           
             14 
             20 
             DNA 
             Artificial sequence 
             
               misc_feature 
               (1)..(20) 
               Primer 
             
           
            14
tctgtaccct gaccttgttg                                                 20
 
           
             15 
             20 
             DNA 
             Artificial sequence 
             
               misc_feature 
               (1)..(20) 
               Primer 
             
           
            15
cgtggatacc acatcgtgat                                                 20
 
           
             16 
             19 
             DNA 
             Artificial sequence 
             
               misc_feature 
               (1)..(19) 
               Primer 
             
           
            16
accttggctt gaagtactc                                                  19
 
           
             17 
             23 
             DNA 
             Artificial sequence 
             
               misc_feature 
               (1)..(23) 
               Primer 
             
           
            17
tcaacctaca gctgatttgg acc                                             23
 
           
             18 
             22 
             DNA 
             Artificial sequence 
             
               misc_feature 
               (1)..(22) 
               Primer 
             
           
            18
ggaccggaac gacaatctga tc                                              22
 
           
             19 
             22 
             DNA 
             Artificial sequence 
             
               misc_feature 
               (1)..(22) 
               Primer 
             
           
            19
ctagggaagt ccaaatcagc cg                                              22
 
           
             20 
             23 
             DNA 
             Artificial sequence 
             
               misc_feature 
               (1)..(23) 
               Primer 
             
           
            20
tttggaccgg aactttccag aag                                             23
 
           
             21 
             23 
             DNA 
             Artificial sequence 
             
               misc_feature 
               (1)..(23) 
               Primer 
             
           
            21
ctaacttgcg ccatcggaga aac                                             23
 
           
             22 
             23 
             DNA 
             Artificial sequence 
             
               misc_feature 
               (1)..(23) 
               Primer 
             
           
            22
gacttgtcac ctggaatacg gac                                             23
 
           
             23 
             23 
             DNA 
             Artificial sequence 
             
               misc_feature 
               (1)..(23) 
               Primer 
             
           
            23
attcttgagc tcatcaagca gcc                                             23
 
           
             24 
             22 
             DNA 
             Artificial sequence 
             
               misc_feature 
               (1)..(22) 
               Primer 
             
           
            24
aaggttggta tcgctggagc tg                                              22
 
           
             25 
             23 
             DNA 
             Artificial sequence 
             
               misc_feature 
               (1)..(23) 
               Primer 
             
           
            25
tctccacaat ggcttcctct atg                                             23
 
           
             26 
             671 
             DNA 
             Artificial sequence 
             
               misc_feature 
               (1)..(671) 
               Probe 
             
           
            26
caagatggat tgcacgcagg ttctccggcc gcttgggtgg agaggctatt cggctatgac     60
tgggcacaac agacaatcgg ctgctctgat gccgccgtgt tccggctgtc agcgcagggg    120
cgcccggttc tttttgtcaa gaccgacctg tccggtgccc tgaatgaact gcaggacgag    180
gcagcgcggc tatcgtggct ggccacgacg ggcgttcctt gcgcagctgt gctcgacgtt    240
gtcactgaag cgggaaggga ctggctgcta ttgggcgaag tgccggggca ggatctcctg    300
tcatctcacc ttgctcctgc cgagaaagta tccatcatgg ctgatgcaat gcggcggctg    360
catacgcttg atccggctac ctgcccattc gaccaccaag cgaaacatcg catcgagcga    420
gcacgtactc ggatggaagc cggtcttgtc gatcaggatg atctggacga agagcatcag    480
gggctcgcgc cagccgaact gttcgccagg ctcaaggcgc gcatgcccga cggcgaggat    540
ctcgtcgtga cccatggcga tgcctgcttg ccgaatatca tggtggaaaa tggccgcttt    600
tctggattca tcgactgtgg ccggctgggt gtggcggacc gctatcagga catagcgttg    660
gctacccgtg a                                                         671
 
           
             27 
             739 
             DNA 
             Artificial sequence 
             
               misc_feature 
               (1)..(739) 
               Amplified fragment 
             
           
            27
ggaccggaac gacaatctga tccccatcaa gcttgagctc aggatttagc agcattccag     60
attgggttca atcaacaagg tacgagccat atcactttat tcaaattggt atcgccaaaa    120
ccaagaagga actcccatcc tcaaaggttt gtaaggaaga attctcagtc caaagcctca    180
acaaggtcag ggtacagagt ctccaaacca ttagccaaaa gctacaggag atcaatgaag    240
aatcttcaat caaagtaaac tactgttcca gcacatgcat catggtcagt aagtttcaga    300
aaaagacatc caccgaagac ttaaagttag tgggcatctt tgaaagtaat cttgtcaaca    360
tcgagcagct ggcttgtggg gaccagacaa aaaaggaatg gtgcagaatt gttaggcgca    420
cctaccaaaa gcatctttgc ctttattgca aagataaagc agattcctct agtacaagtg    480
gggaacaaaa taacgtggaa aagagctgtc ctgacagccc actcactaat gcgtatgacg    540
aacgcagtga cgaccacaaa agaattccct ctatataaga aggcattcat tcccatttga    600
aggatcatca gatactgaac caatccttct agaagatcta agcttatcga taagcttgat    660
gtaattggag gaagatcaaa attttcaatc cccattcttc gattgcttca attgaagttt    720
ctccgatggc gcaagttag                                                 739
 
           
             28 
             757 
             DNA 
             Artificial sequence 
             
               misc_feature 
               (1)..(757) 
               Amplified fragment 
             
           
            28
tcaacctaca gctgatttgg accggaacga caatctgatc cccatcaagc ttgagctcag     60
gatttagcag cattccagat tgggttcaat caacaaggta cgagccatat cactttattc    120
aaattggtat cgccaaaacc aagaaggaac tcccatcctc aaaggtttgt aaggaagaat    180
tctcagtcca aagcctcaac aaggtcaggg tacagagtct ccaaaccatt agccaaaagc    240
tacaggagat caatgaagaa tcttcaatca aagtaaacta ctgttccagc acatgcatca    300
tggtcagtaa gtttcagaaa aagacatcca ccgaagactt aaagttagtg ggcatctttg    360
aaagtaatct tgtcaacatc gagcagctgg cttgtgggga ccagacaaaa aaggaatggt    420
gcagaattgt taggcgcacc taccaaaagc atctttgcct ttattgcaaa gataaagcag    480
attcctctag tacaagtggg gaacaaaata acgtggaaaa gagctgtcct gacagcccac    540
tcactaatgc gtatgacgaa cgcagtgacg accacaaaag aattccctct atataagaag    600
gcattcattc ccatttgaag gatcatcaga tactgaacca atccttctag aagatctaag    660
cttatcgata agcttgatgt aattggagga agatcaaaat tttcaatccc cattcttcga    720
ttgcttcaat tgaagtttct ccgatggcgc aagttag                             757
 
           
             29 
             1057 
             DNA 
             Artificial sequence 
             
               misc_feature 
               (1)..(1057) 
               Amplified fragment 
             
           
            29
tcaacctaca gctgatttgg accggaacga caatctgatc cccatcaagc ttgagctcag     60
gatttagcag cattccagat tgggttcaat caacaaggta cgagccatat cactttattc    120
aaattggtat cgccaaaacc aagaaggaac tcccatcctc aaaggtttgt aaggaagaat    180
tctcagtcca aagcctcaac aaggtcaggg tacagagtct ccaaaccatt agccaaaagc    240
tacaggagat caatgaagaa tcttcaatca aagtaaacta ctgttccagc acatgcatca    300
tggtcagtaa gtttcagaaa aagacatcca ccgaagactt aaagttagtg ggcatctttg    360
aaagtaatct tgtcaacatc gagcagctgg cttgtgggga ccagacaaaa aaggaatggt    420
gcagaattgt taggcgcacc taccaaaagc atctttgcct ttattgcaaa gataaagcag    480
attcctctag tacaagtggg gaacaaaata acgtggaaaa gagctgtcct gacagcccac    540
tcactaatgc gtatgacgaa cgcagtgacg accacaaaag aattccctct atataagaag    600
gcattcattc ccatttgaag gatcatcaga tactgaacca atccttctag aagatctaag    660
cttatcgata agcttgatgt aattggagga agatcaaaat tttcaatccc cattcttcga    720
ttgcttcaat tgaagtttct ccgatggcgc aagttagcag aatctgcaat ggtgtgcaga    780
acccatctct tatctccaat ctctcgaaat ccagtcaacg caaatctccc ttatcggttt    840
ctctgaagac gcagcagcat ccacgagctt atccgatttc gtcgtcgtgg ggattgaaga    900
agagtgggat gacgttaatt ggctctgagc ttcgtcctct taaggtcatg tcttctgttt    960
ccacggcgtg catgcttcac ggtgcaagca gccgtccagc aactgctcgt aagtcctctg   1020
gtctttctgg aaccgtccgt attccaggtg acaagtc                            1057
 
           
             30 
             2352 
             DNA 
             Artificial sequence 
             
               misc_feature 
               (1)..(2352) 
               Amplified fragment 
             
           
            30
tcaacctaca gctgatttgg accggaacga caatctgatc cccatcaagc ttgagctcag     60
gatttagcag cattccagat tgggttcaat caacaaggta cgagccatat cactttattc    120
aaattggtat cgccaaaacc aagaaggaac tcccatcctc aaaggtttgt aaggaagaat    180
tctcagtcca aagcctcaac aaggtcaggg tacagagtct ccaaaccatt agccaaaagc    240
tacaggagat caatgaagaa tcttcaatca aagtaaacta ctgttccagc acatgcatca    300
tggtcagtaa gtttcagaaa aagacatcca ccgaagactt aaagttagtg ggcatctttg    360
aaagtaatct tgtcaacatc gagcagctgg cttgtgggga ccagacaaaa aaggaatggt    420
gcagaattgt taggcgcacc taccaaaagc atctttgcct ttattgcaaa gataaagcag    480
attcctctag tacaagtggg gaacaaaata acgtggaaaa gagctgtcct gacagcccac    540
tcactaatgc gtatgacgaa cgcagtgacg accacaaaag aattccctct atataagaag    600
gcattcattc ccatttgaag gatcatcaga tactgaacca atccttctag aagatctaag    660
cttatcgata agcttgatgt aattggagga agatcaaaat tttcaatccc cattcttcga    720
ttgcttcaat tgaagtttct ccgatggcgc aagttagcag aatctgcaat ggtgtgcaga    780
acccatctct tatctccaat ctctcgaaat ccagtcaacg caaatctccc ttatcggttt    840
ctctgaagac gcagcagcat ccacgagctt atccgatttc gtcgtcgtgg ggattgaaga    900
agagtgggat gacgttaatt ggctctgagc ttcgtcctct taaggtcatg tcttctgttt    960
ccacggcgtg catgcttcac ggtgcaagca gccgtccagc aactgctcgt aagtcctctg   1020
gtctttctgg aaccgtccgt attccaggtg acaagtctat ctcccacagg tccttcatgt   1080
ttggaggtct cgctagcggt gaaactcgta tcaccggtct tttggaaggt gaagatgtta   1140
tcaacactgg taaggctatg caagctatgg gtgccagaat ccgtaaggaa ggtgatactt   1200
ggatcattga tggtgttggt aacggtggac tccttgctcc tgaggctcct ctcgatttcg   1260
gtaacgctgc aactggttgc cgtttgacta tgggtcttgt tggtgtttac gatttcgata   1320
gcactttcat tggtgacgct tctctcacta agcgtccaat gggtcgtgtg ttgaacccac   1380
ttcgcgaaat gggtgtgcag gtgaagtctg aagacggtga tcgtcttcca gttaccttgc   1440
gtggaccaaa gactccaacg ccaatcacct acagggtacc tatggcttcc gctcaagtga   1500
agtccgctgt tctgcttgct ggtctcaaca ccccaggtat caccactgtt atcgagccaa   1560
tcatgactcg tgaccacact gaaaagatgc ttcaaggttt tggtgctaac cttaccgttg   1620
agactgatgc tgacggtgtg cgtaccatcc gtcttgaagg tcgtggtaag ctcaccggtc   1680
aagtgattga tgttccaggt gatccatcct ctactgcttt cccattggtt gctgccttgc   1740
ttgttccagg ttccgacgtc accatcctta acgttttgat gaacccaacc cgtactggtc   1800
tcatcttgac tctgcaggaa atgggtgccg acatcgaagt gatcaaccca cgtcttgctg   1860
gtggagaaga cgtggctgac ttgcgtgttc gttcttctac tttgaagggt gttactgttc   1920
cagaagaccg tgctccttct atgatcgacg agtatccaat tctcgctgtt gcagctgcat   1980
tcgctgaagg tgctaccgtt atgaacggtt tggaagaact ccgtgttaag gaaagcgacc   2040
gtctttctgc tgtcgcaaac ggtctcaagc tcaacggtgt tgattgcgat gaaggtgaga   2100
cttctctcgt cgtgcgtggt cgtcctgacg gtaagggtct cggtaacgct tctggagcag   2160
ctgtcgctac ccacctcgat caccgtatcg ctatgagctt cctcgttatg ggtctcgttt   2220
ctgaaaaccc tgttactgtt gatgatgcta ctatgatcgc tactagcttc ccagagttca   2280
tggatttgat ggctggtctt ggagctaaga tcgaactctc cgacactaag gctgcttgat   2340
gagctcaaga at                                                       2352
 
           
             31 
             1039 
             DNA 
             Artificial sequence 
             
               misc_feature 
               (1)..(1039) 
               Amplified fragment 
             
           
            31
ggaccggaac gacaatctga tccccatcaa gcttgagctc aggatttagc agcattccag     60
attgggttca atcaacaagg tacgagccat atcactttat tcaaattggt atcgccaaaa    120
ccaagaagga actcccatcc tcaaaggttt gtaaggaaga attctcagtc caaagcctca    180
acaaggtcag ggtacagagt ctccaaacca ttagccaaaa gctacaggag atcaatgaag    240
aatcttcaat caaagtaaac tactgttcca gcacatgcat catggtcagt aagtttcaga    300
aaaagacatc caccgaagac ttaaagttag tgggcatctt tgaaagtaat cttgtcaaca    360
tcgagcagct ggcttgtggg gaccagacaa aaaaggaatg gtgcagaatt gttaggcgca    420
cctaccaaaa gcatctttgc ctttattgca aagataaagc agattcctct agtacaagtg    480
gggaacaaaa taacgtggaa aagagctgtc ctgacagccc actcactaat gcgtatgacg    540
aacgcagtga cgaccacaaa agaattccct ctatataaga aggcattcat tcccatttga    600
aggatcatca gatactgaac caatccttct agaagatcta agcttatcga taagcttgat    660
gtaattggag gaagatcaaa attttcaatc cccattcttc gattgcttca attgaagttt    720
ctccgatggc gcaagttagc agaatctgca atggtgtgca gaacccatct cttatctcca    780
atctctcgaa atccagtcaa cgcaaatctc ccttatcggt ttctctgaag acgcagcagc    840
atccacgagc ttatccgatt tcgtcgtcgt ggggattgaa gaagagtggg atgacgttaa    900
ttggctctga gcttcgtcct cttaaggtca tgtcttctgt ttccacggcg tgcatgcttc    960
acggtgcaag cagccgtcca gcaactgctc gtaagtcctc tggtctttct ggaaccgtcc   1020
gtattccagg tgacaagtc                                                1039
 
           
             32 
             2334 
             DNA 
             Artificial sequence 
             
               misc_feature 
               (1)..(2334) 
               Amplified fragment 
             
           
            32
ggaccggaac gacaatctga tccccatcaa gcttgagctc aggatttagc agcattccag     60
attgggttca atcaacaagg tacgagccat atcactttat tcaaattggt atcgccaaaa    120
ccaagaagga actcccatcc tcaaaggttt gtaaggaaga attctcagtc caaagcctca    180
acaaggtcag ggtacagagt ctccaaacca ttagccaaaa gctacaggag atcaatgaag    240
aatcttcaat caaagtaaac tactgttcca gcacatgcat catggtcagt aagtttcaga    300
aaaagacatc caccgaagac ttaaagttag tgggcatctt tgaaagtaat cttgtcaaca    360
tcgagcagct ggcttgtggg gaccagacaa aaaaggaatg gtgcagaatt gttaggcgca    420
cctaccaaaa gcatctttgc ctttattgca aagataaagc agattcctct agtacaagtg    480
gggaacaaaa taacgtggaa aagagctgtc ctgacagccc actcactaat gcgtatgacg    540
aacgcagtga cgaccacaaa agaattccct ctatataaga aggcattcat tcccatttga    600
aggatcatca gatactgaac caatccttct agaagatcta agcttatcga taagcttgat    660
gtaattggag gaagatcaaa attttcaatc cccattcttc gattgcttca attgaagttt    720
ctccgatggc gcaagttagc agaatctgca atggtgtgca gaacccatct cttatctcca    780
atctctcgaa atccagtcaa cgcaaatctc ccttatcggt ttctctgaag acgcagcagc    840
atccacgagc ttatccgatt tcgtcgtcgt ggggattgaa gaagagtggg atgacgttaa    900
ttggctctga gcttcgtcct cttaaggtca tgtcttctgt ttccacggcg tgcatgcttc    960
acggtgcaag cagccgtcca gcaactgctc gtaagtcctc tggtctttct ggaaccgtcc   1020
gtattccagg tgacaagtct atctcccaca ggtccttcat gtttggaggt ctcgctagcg   1080
gtgaaactcg tatcaccggt cttttggaag gtgaagatgt tatcaacact ggtaaggcta   1140
tgcaagctat gggtgccaga atccgtaagg aaggtgatac ttggatcatt gatggtgttg   1200
gtaacggtgg actccttgct cctgaggctc ctctcgattt cggtaacgct gcaactggtt   1260
gccgtttgac tatgggtctt gttggtgttt acgatttcga tagcactttc attggtgacg   1320
cttctctcac taagcgtcca atgggtcgtg tgttgaaccc acttcgcgaa atgggtgtgc   1380
aggtgaagtc tgaagacggt gatcgtcttc cagttacctt gcgtggacca aagactccaa   1440
cgccaatcac ctacagggta cctatggctt ccgctcaagt gaagtccgct gttctgcttg   1500
ctggtctcaa caccccaggt atcaccactg ttatcgagcc aatcatgact cgtgaccaca   1560
ctgaaaagat gcttcaaggt tttggtgcta accttaccgt tgagactgat gctgacggtg   1620
tgcgtaccat ccgtcttgaa ggtcgtggta agctcaccgg tcaagtgatt gatgttccag   1680
gtgatccatc ctctactgct ttcccattgg ttgctgcctt gcttgttcca ggttccgacg   1740
tcaccatcct taacgttttg atgaacccaa cccgtactgg tctcatcttg actctgcagg   1800
aaatgggtgc cgacatcgaa gtgatcaacc cacgtcttgc tggtggagaa gacgtggctg   1860
acttgcgtgt tcgttcttct actttgaagg gtgttactgt tccagaagac cgtgctcctt   1920
ctatgatcga cgagtatcca attctcgctg ttgcagctgc attcgctgaa ggtgctaccg   1980
ttatgaacgg tttggaagaa ctccgtgtta aggaaagcga ccgtctttct gctgtcgcaa   2040
acggtctcaa gctcaacggt gttgattgcg atgaaggtga gacttctctc gtcgtgcgtg   2100
gtcgtcctga cggtaagggt ctcggtaacg cttctggagc agctgtcgct acccacctcg   2160
atcaccgtat cgctatgagc ttcctcgtta tgggtctcgt ttctgaaaac cctgttactg   2220
ttgatgatgc tactatgatc gctactagct tcccagagtt catggatttg atggctggtc   2280
ttggagctaa gatcgaactc tccgacacta aggctgcttg atgagctcaa gaat         2334
 
           
             33 
             934 
             DNA 
             Artificial sequence 
             
               misc_feature 
               (1)..(934) 
               Amplified fragment 
             
           
            33
aaggttggta tcgctggagc tggaatcgtt ggtgtttgca ctgctttgat gcttcaacgt     60
cgtggattca aggttacctt gattgatcca aacccaccag gtgaaggtgc ttctttcggt    120
aacgctggtt gcttcaacgg ttcctccgtt gttccaatgt ccatgccagg aaacttgact    180
agcgttccaa agtggcttct tgacccaatg ggtccattgt ccatccgttt cagctacttt    240
ccaaccatca tgccttggtt gattcgtttc ttgcttgctg gaagaccaaa caaggtgaag    300
gagcaagcta aggcactccg taacctcatc aagtccactg tgcctttgat caagtccttg    360
gctgaggagg ctgatgctag ccaccttatc cgtcacgaag gtcaccttac cgtgtaccgt    420
ggagaagcag acttcgccaa ggaccgtgga ggttgggaac ttcgtcgtct caacggtgtt    480
cgtactcaaa tcctcagcgc tgatgcattg cgtgatttcg atcctaactt gtctcacgcc    540
tttaccaagg gaatccttat cgaagagaac ggtcacacca tcaacccaca aggtctcgtg    600
actctcttgt ttcgtcgttt catcgctaac ggtggagagt tcgtgtctgc tcgtgttatc    660
ggattcgaga ctgaaggtcg tgctctcaag ggtatcacca ccaccaacgg tgttcttgct    720
gttgatgcag ctgttgttgc agctggtgca cactccaagt ctcttgctaa ctcccttggt    780
gatgacatcc cattggatac cgaacgtgga taccacatcg tgatcgccaa cccagaagct    840
gctccacgta ttccaactac cgatgcttct ggaaagttcc ggtccaaatt tgtttacatt    900
gtgtccaaat ttcggctgat ttggacttcc ctag                                934
 
           
             34 
             1224 
             DNA 
             Artificial sequence 
             
               misc_feature 
               (1)..(1224) 
               Amplified fragment 
             
           
            34
tctccacaat ggcttcctct atgctctctt ccgctactat ggttgcctct ccggctcagg     60
ccactatggt cgctcctttc aacggactta agtcctccgc tgccttccca gccacccgca    120
aggctaacaa cgacattact tccatcacaa gcaacggcgg aagagttaac tgcatgcagg    180
tgtggcctcc gattggaaag aagaagtttg agactctctc ttaccttcct gaccttaccg    240
attccggtgg tcgcgtcaac tgcatgcagg ccatggctga gaaccacaag aaggttggta    300
tcgctggagc tggaatcgtt ggtgtttgca ctgctttgat gcttcaacgt cgtggattca    360
aggttacctt gattgatcca aacccaccag gtgaaggtgc ttctttcggt aacgctggtt    420
gcttcaacgg ttcctccgtt gttccaatgt ccatgccagg aaacttgact agcgttccaa    480
agtggcttct tgacccaatg ggtccattgt ccatccgttt cagctacttt ccaaccatca    540
tgccttggtt gattcgtttc ttgcttgctg gaagaccaaa caaggtgaag gagcaagcta    600
aggcactccg taacctcatc aagtccactg tgcctttgat caagtccttg gctgaggagg    660
ctgatgctag ccaccttatc cgtcacgaag gtcaccttac cgtgtaccgt ggagaagcag    720
acttcgccaa ggaccgtgga ggttgggaac ttcgtcgtct caacggtgtt cgtactcaaa    780
tcctcagcgc tgatgcattg cgtgatttcg atcctaactt gtctcacgcc tttaccaagg    840
gaatccttat cgaagagaac ggtcacacca tcaacccaca aggtctcgtg actctcttgt    900
ttcgtcgttt catcgctaac ggtggagagt tcgtgtctgc tcgtgttatc ggattcgaga    960
ctgaaggtcg tgctctcaag ggtatcacca ccaccaacgg tgttcttgct gttgatgcag   1020
ctgttgttgc agctggtgca cactccaagt ctcttgctaa ctcccttggt gatgacatcc   1080
cattggatac cgaacgtgga taccacatcg tgatcgccaa cccagaagct gctccacgta   1140
ttccaactac cgatgcttct ggaaagttcc ggtccaaatt tgtttacatt gtgtccaaat   1200
ttcggctgat ttggacttcc ctag                                          1224
 
           
             35 
             888 
             DNA 
             Artificial sequence 
             
               misc_feature 
               (1)..(888) 
               Amplified fragment 
             
           
            35
aaggttggta tcgctggagc tggaatcgtt ggtgtttgca ctgctttgat gcttcaacgt     60
cgtggattca aggttacctt gattgatcca aacccaccag gtgaaggtgc ttctttcggt    120
aacgctggtt gcttcaacgg ttcctccgtt gttccaatgt ccatgccagg aaacttgact    180
agcgttccaa agtggcttct tgacccaatg ggtccattgt ccatccgttt cagctacttt    240
ccaaccatca tgccttggtt gattcgtttc ttgcttgctg gaagaccaaa caaggtgaag    300
gagcaagcta aggcactccg taacctcatc aagtccactg tgcctttgat caagtccttg    360
gctgaggagg ctgatgctag ccaccttatc cgtcacgaag gtcaccttac cgtgtaccgt    420
ggagaagcag acttcgccaa ggaccgtgga ggttgggaac ttcgtcgtct caacggtgtt    480
cgtactcaaa tcctcagcgc tgatgcattg cgtgatttcg atcctaactt gtctcacgcc    540
tttaccaagg gaatccttat cgaagagaac ggtcacacca tcaacccaca aggtctcgtg    600
actctcttgt ttcgtcgttt catcgctaac ggtggagagt tcgtgtctgc tcgtgttatc    660
ggattcgaga ctgaaggtcg tgctctcaag ggtatcacca ccaccaacgg tgttcttgct    720
gttgatgcag ctgttgttgc agctggtgca cactccaagt ctcttgctaa ctcccttggt    780
gatgacatcc cattggatac cgaacgtgga taccacatcg tgatcgccaa cccagaagct    840
gctccacgta ttccaactac cgatgcttct ggaaagttcc ggtccaaa                 888
 
           
             36 
             1178 
             DNA 
             Artificial sequence 
             
               misc_feature 
               (1)..(1178) 
               Amplified fragment 
             
           
            36
tctccacaat ggcttcctct atgctctctt ccgctactat ggttgcctct ccggctcagg     60
ccactatggt cgctcctttc aacggactta agtcctccgc tgccttccca gccacccgca    120
aggctaacaa cgacattact tccatcacaa gcaacggcgg aagagttaac tgcatgcagg    180
tgtggcctcc gattggaaag aagaagtttg agactctctc ttaccttcct gaccttaccg    240
attccggtgg tcgcgtcaac tgcatgcagg ccatggctga gaaccacaag aaggttggta    300
tcgctggagc tggaatcgtt ggtgtttgca ctgctttgat gcttcaacgt cgtggattca    360
aggttacctt gattgatcca aacccaccag gtgaaggtgc ttctttcggt aacgctggtt    420
gcttcaacgg ttcctccgtt gttccaatgt ccatgccagg aaacttgact agcgttccaa    480
agtggcttct tgacccaatg ggtccattgt ccatccgttt cagctacttt ccaaccatca    540
tgccttggtt gattcgtttc ttgcttgctg gaagaccaaa caaggtgaag gagcaagcta    600
aggcactccg taacctcatc aagtccactg tgcctttgat caagtccttg gctgaggagg    660
ctgatgctag ccaccttatc cgtcacgaag gtcaccttac cgtgtaccgt ggagaagcag    720
acttcgccaa ggaccgtgga ggttgggaac ttcgtcgtct caacggtgtt cgtactcaaa    780
tcctcagcgc tgatgcattg cgtgatttcg atcctaactt gtctcacgcc tttaccaagg    840
gaatccttat cgaagagaac ggtcacacca tcaacccaca aggtctcgtg actctcttgt    900
ttcgtcgttt catcgctaac ggtggagagt tcgtgtctgc tcgtgttatc ggattcgaga    960
ctgaaggtcg tgctctcaag ggtatcacca ccaccaacgg tgttcttgct gttgatgcag   1020
ctgttgttgc agctggtgca cactccaagt ctcttgctaa ctcccttggt gatgacatcc   1080
cattggatac cgaacgtgga taccacatcg tgatcgccaa cccagaagct gctccacgta   1140
ttccaactac cgatgcttct ggaaagttcc ggtccaaa                           1178