Genetic Classification And Cloning Of Transparent Testa Homologous Genes In Brassica Napus L.

Oilseed rape is the main oil crop and the most important cultivated type in our country. Owing to thinner hull, less fiber, higher oil and protein content than its black-seeded counterpart with a dark, thicker, and more lignified seed coat, yellow-seeded variety is an important goal for breeding of oilseed Brassica napus. There are many stability of yellow-seeded genotypes in B.rapa and B.juncea, many genes have been imported into the yellow-seeded of Brassica napus, formed a relatively abundant genetic diversity of special groups. The yellow-seeded variety of Brassica napus is widespread " not yellow enough" problem, and it’s difficult to improve the stability, penetrance and degree of yellow-seeded. Its genetic model is more complicated.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, known as the Transparent Testa mutations in Arabidopsis. 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.We tested the allelisms among different yellow-seeded accessions and studied the genetic diversity through clustering. The results could provide the theoretical basis for the heterosis utilization of yellow-seeded rapeseed. At the same time, we designed primers according to Transparent Testa genes of Arabidopsis in the area of the conservative. The yellow-seeded Youyan10 and black-seeded G142 of Brassica napus were used for PCR amplification and cloning sequencing. The main results were as follows:1.According to the result of allelism test, by investigation of the segregation of seed coat colour in F1 and F2 of cross between different accessions, 11 yellow-seeded rapeseed accessions can be divided into five groups: group I including Youyan10, CZV55, E718, and Arm, mostly followed a dominant inheritance model; group II including Q33 and D615, in incomplete dominant inheritance; group III including 2006 C, X2006, and 740C; the rest two groups only including Polo and HY15, respectively. The last three groups were generally in recessive genetic model. However, the inheritance of yellow-seeded traits of Youyan10 and 2006 C was various from dominant to recessive when they crossed with the different genotypes of brown-seeded.2.Cluster based on SSR molecular polymorphism showed that 35 yellow-seeded rapeseed accessions could be classified into three main groups: B.napus, B.rapa, and B.juncea. The group B.napus could be further divided into eight subgroups, the representations of them were 2006 C and 740 C, Youyan system, Q33, Ramiro from French, GQ4 from Shaanxi, Arm and prof from Canada, and yellow Polo from Polish cultivar Polo. This generally agrees with the pedigrees, breeders and originations.3.The full-length genomic sequence of BnTT6 is 1631 bp with three exons and two introns. Two introns are in sequence of 397-744 bp and 1174-1262 bp, respectively. TT6 genetic structure of Brassica napus and Arabidopsis thaliana were linear highly consistent. BnTT6 protein contains 358 amino acid residues, and share 93.1% identity with AtTT6. The full-length genomic sequence of BnTT7 is 3038 bp with four exons and three introns. Three introns are in sequence of 465-1533 bp, 1981-2063 bp and 2234-2303 bp, respectively. TT7 genetic structure of Brassica napus and Arabidopsis thaliana were linear highly consistent. BnTT7 protein contains 511 amino acid residues, and share 88.9% identity with AtTT7. The sequence of BnTT19 is 986 bp with three exons and two introns. Two introns are in sequence of 206-291 bp and 340-414 bp, respectively. TT19 genetic structure of Brassica napus and Arabidopsis thaliana were linear highly consistent. BnTT19 protein contains 215 amino acid residues, and share 85.0% identity with AtTT19.