| Seedless grapes have high commercial value and seedlessness for table grapes and raisins is one of the principle objectives of Vitis vinifera grape breeders. Recent studies have shown that the seedless trait in grapes is controlled by one major and some minor genes. The position and mapping of the major gene for seedlessness was studied in this research using Red Globe, Flame Seedless, and Thompson Seedless, together with Fl progenies from Red Globe X Flame Seedless crosses. The sequences of marker linked to the major seedlessness gene were analyzed by BLAST in GenBank and the open reading frame (ORF) of these sequences was also found in GenBank. 39970524-5-564 DNA labeled with a DIG DNA Labeling and Detection Kit was used as a probe against Southern blot with genomic DNA from grapes. The main results were as follows.1. Identification of SCAR markers linked to grape seedlessness geneNine primers (including UBC-269 and GSLP1) were designed and synthesized based on DNA sequences of UBC-2694g4 and GSLP1569. The template DNA from Red Globe (seeded paternal parent) and Flame Seedless (seedless maternal parent) were screened using these primers. For Flame Seedless, GSLP1 yielded specific marker GSLPlseg; No.39970524-5 primer yielded specific marker 39970524-5-564; and No.6 primer yielded specific marker 39970524-6-1538 and 39970524-6-1200. GSLP1, No. 39970524-5, and No. 39970524-6 primers were used specifically to screen template DNA from the experimental plant materials: results showed that the specific markers GSLPlseg, 39970524-5-564, 39970524-6-1538 and 39970524-6-1200 were co-segregating with the major seedlessness gene. All these specific loci were also present in Thompson Seedless which was the initial donor of the seedlessness gene. It suggests that these SCAR markers are linked to a major grape seedlessness gene S .2. Position and mapping for grape seedlessness geneMarker order and map distance were estimated using the software 'QTXbl7'. This showed that GSLP1569, 39970524-5-564, 39970524-6-1538 and 39970524-6-1200 wereIVtightly linked to gene S. When P=0.01, confidence limits for map distance ranged from 0.2 to 9.9; standard errors of map distance were from 0.6 to 1.9; LOD for linkage were from 32.7 to 46.4. These markers and the gene S were found to be in the same group. The markers were either sides of gene S, covering 12.3 cM of the grape genome. The genetic distances between gene S and 39970524-5-564, GSLP1569, 39970524-6-1538 and 39970524-6-1200 were 0.6 cM, 1.2 cM, 4.9 cM and 11.1 cM respectively.3. Comparative analysis of the DNA sequences of specific markers linked to grape seedlessness geneDNA fragments of 39970524-5-564 and 39970524-6-1538 derived from Flame Seedless were retrieved from agarose gel and ligated into the pGEMB-T Easy Vector System, and then transformed into E. coli DH5-a. Sequencing of positive clones was carried out and the result showed that the 39970524-5-564 was 564bp and the 39970524-6-1538 was 1538bp. The sequences data had submitted to GenBank. The GenBank accession number for 39970524-5-564 is AY327513 and 39970524-6-1538 is AY327514.It has been suggested that the sequences of 39970524-5-564 and 39970524-6-1538 are unique to the grape (Vitis viniferd) genome. But, these sequences were extensively compared by BLAST in GenBank and a small part was found to be homologous. 22 base pairs from the sequence of 39970524-5-564 (from 38 to 59) matched two nucleic acid sequences coded MAP3P from Arabidopsis thaliana and also matched the sequence for Arabidopsis thaliana BAG T17H7 from Chromosome 1. Further study is needed to determine the significance of the 22 base pairs from the sequence of 39970524-5-564. The sequence of 39970524-6-1538 was not homologous with Arabidopsis thaliana, but it matched rice [Oryza sativa (Japonica cvs)] genomic DNA from chromosome 6.4. Finding ORF's of the DNA sequences of specific markers linked to grape seedlessness geneORFs of the sequences of markers linked to gene S were found in GenBank. There was an ORF in the sequen... |
