Analysis Of Transcriptome And Genome-wide DNA Methylation In Yellow Seed Coat Of Brassica Napus

Rape has long been the main oil crop in southern my country and occupies an important position in my country’s agricultural production.The seed coat color of rape is an important agronomic trait.The seeds of yellow-seed rape have the characteristics of high oil and protein content and low fiber content,which have great advantages in the subsequent production and processing of rape.However,there is no natural yellow-seed mutant in Brassica oleracea,and the existing Brassica napus yellow-seed rape is obtained by crossing with other varieties,and usually has the problem of unstable characters.The origin of the traits of rape yellow seed is generally considered to be the lack of flavonols and procyanidins in the seed coat of flavonoid synthesis pathway products.The proanthocyanidins synthesized by the flavonoid synthesis pathway undergo oxidation to synthesize anthocyanins,which makes the seed coat appear brown or black.The precursor phenolic Co A in the flavonoid synthesis pathway is derived from the phenylpropanoid pathway.In order to analyze the reasons for the genetic instability of the yellow seed trait from the genetic level,this study used the genetically stable yellow seed trait Huang Ai Zao and the yellow seed trait genetically unstable Huahuang 1 and GH06 as the materials.Transcriptome sequencing and whole-genome DNA methylation sequencing analysis were performed.The main findings are as follows:(1)The expression of flavonoid synthesis pathway genes Bn FLS2,Bn FL3 c,Bn MYB5 b and Bn TT6 b decreased in seed coat of Huang Ai Zao.Transcriptome analysis of the seed coats of three cultivars at 15,25 and 35 days after pollination revealed that seven flavonoid synthesis-related genes Bn FL3 c,Bn TT6 b,Bn PAL4 a,Bn TT18 c,Bn TT19 b,Bn MYB5 B and Bn FLS2 were found in Huangaizao and Huaihuai.There are differences between Hua Huang 1 and GH06.Among them,the expression levels of Bn FLS2,Bn FL3 c,Bn MYB5 B and Bn TT6 b decreased in the seed coat of Huang Ai Zao at different developmental stages;the expression levels of Bn PAL4 a,Bn TT18 c and Bn TT19 b increased in the different development stages of Huang Ai Zao.(2)The expression levels of DNA methyltransferase gene Bn DRM1 b and demethylase gene Bn DML4 a increased in the seed coat of Huang Ai Zao,and another demethylase gene Bn DML5 b was expressed in the seed coat of Huang Ai Zao at different developmental stages expression decreased.The expression levels of genes Bn CPSF30 a,Bn CPSF30 e and Bn ECT2 c encoding three different m6 A reading proteins were significantly higher than those of GH06 and Huahuang 1 in seed coat of Huang Ai Zao(3)The methylation levels of the flavonoid synthesis pathway genes Bn FL3 c and Bn TT6 b were increased in the seed coat of Huang Ai Zao.By comparing the wholegenome DNA methylation of the seed coats of the three varieties at different developmental stages,it was found that the whole-genome DNA methylation level of Huang’aizao was higher than that of Huahuang 1 and GH06.It was found that the DNA methylation levels of Bn FL3 c and Bn TT6 b in the seed coat of Huang Ai Zao at15 and 35 days after pollination were significantly higher than those of GH06 and Huahuang 1.The methylation of specific elements was analyzed,and it was found that Bn FL3 c mainly had CG hypermethylation in the downstream region of the gene.Bn TT6 b has CHH hypermethylation in the promoter region,and CG and CHG hypermethylation in the exon and intron regions,respectively.Therefore,it is speculated that these two genes may reduce their expression levels due to DNA hypermethylation,thus making the yellow seed traits of Huang Aizao stable.(4)The Huang Ai Zao transposon showed the characteristics of hypermethylation.According to the overall hypermethylation characteristics of Huang’aizao’s genome,we analyzed the expression levels of repetitive sequences in the samples,and found that the expression level of the LTR retrotransposon,the main component of retrotransposons,was significantly higher in GH06 than in Huang’aizao and Huahuang.Number one.The expression levels of LTR retrotransposon in Huahuang No.1 and Huang’aizao were basically the same.The expression levels of most nonLTR retrotransposons in Huangaizao were significantly higher than those in GH06 and Huahuang 1,such as-CTA and-TGG families,while the non-LTR transposons between Huahuang 1 and GH06 were reversed.There was no significant difference in the expression levels of transposons.Analysis of methylation 3 in the transposon region of the genome showed that the methylation level of the transposon in Huahuang 1 was higher than that in Huang’aizao,and the methylation level of Huang’aizao was higher than that in GH06.This provides clues for the future in-depth study of the regulatory mechanism of the genetic stability of yellow seed traits.