Abstract:

Seed coat color is not only a morphological indicator but also association with dormancy and quality of seed in Braasica. The seeds of Brassica species are divided into such categories as black, brown or yellow according to their coat color. The emphasis has been put on breeding for the yellow-seeded rapeseed in recent years. Most genetic studies showed that seed coat color is controlled by two duplicate loci in Brassica juncea. Chemical analysis and DMACA (p-dimethylaminocinnamaldehyde) staining confirmed that the yellow seed coat does not contain any proanthocyanidins while the black or brown seed coat does. Dihydroflavonol 4-reductase (DFR), anthocyanidin synthase (ANS), and anthocyanidin reductase (ANR) are the key enzymes of flavonoid biosynthetic pathway leading to production of proanthocyanidins. As revealed by RT-PCR, the genes encoding DFR, ANS and ANR are not expressed in the transparent testa of B. juncea yellow seeds. The two BC₆F₂ populations, derived from crossing and backcrossing the yellow-seeded parent Sichuan yellow to the black-seeded Purple-leaf mustard and comprised 143 or 141 individual plants, were used to map the loci controlling seed coat color in B. juncea by using SSRs and SCARs. The twenty-two B. juncea accessions collected from all over the world were genotyped for association analysis using the markers mapped. The expression of the genes encoding DFR, ANS, and ANR in seed coat at 20 days after pollination was investigated by RT-PCR in the twelve accessions. The allelism test was carried out by a diallel cross for the six yellow-seeded accessions from China, Canada, India or Russia. We mapped the two loci A and B controlling seed coat color on the linkage groups A9 and B3, receptively, and found the 22 linked markers flanking these loci. The closest markers flanking the locus A on A9 were the co-dominant markers SCM08 and Ni4-C09, which are 0.5 and 1.6 cM far from the locus, respectively. Although the marker CB10298 on one side of the locus B on B3 was also a co-dominant SSR marker with 0.8 cM from locus B, on the other side a RAPD marker S1096_-700 with 3.3 cM apart. The 0.9 and 1.5 cM-long chromosomal regions around these loci were revealed by association analysis to be conserved in all the yellow-seeded accessions genotyped. The expression of the genes DFR and ANS was not detected by RT-PCR in the yellow-seeded accessions investigated although these genes were found to be expressed in seed coat of all the black-seeded ones. The gene ANR was strongly expressed in all the black-seeded accessions studied, and not or weakly expressed in the yellow-seeded accessions. The F₁ plants of the fifteen combinations from diallel crossing of the six yellow-seeded accessions all produced yellow seeds, indicating the loci for seed coat color were allelic to each other in these parents. It is proposed from these results and previous studies that the gene controlling seed coat color is a transcription-factor-coding gene and all the yellow-seeded accessions derive from a single origin in Brassica juncea.

Key words: Brassica Juncea, Seed coat color, Genetic mapping, Association analysis, Allelism, Gene expression