Construction Of Core Collections Of Yellow-seeded Rapeseed(Brassica Napus L) And The Screening Of Related Single Nucleotide Polymorphism Markers

Oilseed rape is the main oil crop and the most important cultivated type in our country. Compared to black-seeded rapeseed the yellow-seeded rapeseed has a varieties of advantages that which is thinner seed coat, less pigments, more proteins and less lignose in the same background. Because the natural yellow-seeded rapeseed doesn’t exit, the genetic background of yellow-seeded rapeseed is limited, so the core germplasm construction of yellow-seeded rapeseed will make full use of germplasm resources and speed up breeding process.The anatomical structure of seedcoat growth and pigment biosynthesis pathway in Arabidopisis have been deeply studied, which have important guiding significance for explaining synthesis mechanism of seedcoat colour from the molecular level.Mutations of proteins gene and some key enzymes encoding flavonoids biosynthesis cause changes in flavonoids accumulation or the biosynthesis of final products, and result in seedcoat color variation to different extents. Brassica and Arabidopsis are both the members of Cruciferae, and their sequence conservation reaches 86%. The change of seedcoat color and pigments biosynthesis mechanism of Brassica are similar to Arabidopsis. The researches on flavonoids biosynthesis of the related gene in Arabidopsis have guiding significance to the formation mechanism of yellow seedcoat in yellow-seeded rapeseed.The paper used SSR markers to analysise the genetic diversity of 180 parental germplasm of yellow-seeded B. napus provided by the Chongqing Rapeseed Engineering Research Center of Southwest University. Then the core collection of yellow-seeded rapeseed (Brassica napus L) was constructed based on the cluster analysis results and material character data.The research used homology-based cloning (?)y, and the primers were designed basing on conservative sequence of the related transparent seed coat gene between Brassica napus L and the model plant Arabidopsis thaliana. The gene fragments were sequenced.It was expected to find the single nuclcotide mutations for seedcoat color and translate into function marker, being used in molecular marker assisted breeding. The results were as follows:1. Analysis of genetic diversity of yellow-seeded rapeseed (Brassica napus L)The genetic diversity of 180 parental germplasm of yellow-seeded B. napus, which were provided by the Chongqing Rapeseed Engineering Research Center of Southwest University, was analyzed.308 alleles were detected via 60 SSR primer pairs which were selected from the different linked groups in Brasicca napus.207 polymorphic bands were obtained, accounting for 67.2%of the total bands. The UPGMA cluster analysis basing on SSR data showed that, the genetic similarity coefficient was 0.56-1.00 and 180 parental lines could be classified into three groups at the threshold 0.566 of genetic distance, which agreeed with the pedigree analysis.2. Constrcution of core collections of 180 parental germplasm f yellow-seeded B. napus.20 samples were chosen as core collection according to the pedigree, genetic distance and five quality traits of germplasm characteristics.And the sampling ratio was 11.11%. After comparison, the samples had no significant difference with the primary core collection.So the primary core collection could represent the genetic diversity of the original accessions.3. Screening of the single nucleotide mutations for seedcoat colorThe 19 genes, which were related with transparent seedcoat color, were coloned from yellow-seeded and blacked-seeded rapeseed accessions using homology-based cloning.And 14 genes got the corresponding copy sequence,which there were multiple copies of the gene sequence and single nucleotide polymorphisms sites in.It got the only restriction enzyme site that had effect on the seed coat color from the gene BnTT3, BnTT6 and BnTT18.The amplified productions were restricted using restriction enzyme and it showed that the number of bands wer different from that we predicted.It didn’t confirm that if the mutations of these SNP sites had effect on the seed coat color.It still need more research.