| Glucosinolates is one of important secondary metabolites in cruciferae family,and it is an important nutrient composition in Brassica rapa crops. It will be taketime and energy to directly determine of glucosinolates, which is unfavorableselection directly in the process of breeding. Thus it is necessary to study thevariation in total glucosinolates and different components at the molecular level,helping to further explore the elite genetic resources.In this study, based on genome sequence information of Chinese cabbage,specific SSR loci linked to all genes of glucosinolate synthesis were developed. Therepeat types and distribution characteristics of SSRs were futher analyzed.Combining InDeL markers developed throughout the whole genome level, theamplyfication and polymorphism level in75Chinese cabbage inbred lines withdifferent types were analyzed. By HPLC (high performance liquid chromatography)analysis, glucosinolate level in leaves at seedling stage (35days after tansplanting)and mature period (suitable havest stage) was detected. Through associationmapping analysis, SSR allelic genes or loci and InDeL markers linked toglucosinolate level in leaves were found. This infromation would provide basicmaterials for marker-assisted selection breeding and accelerate the process ofnutritional quality breeding in Chinese cabbage. The main results are listed asfollows:1. Compared with sequences of glucosinolate biosynthesis genes in arabidopsisand Chinese cabbage, we found multiple homologous genes in Chinese cabbage of52glucosinolate biosynthesis genes in Arabidopsis:8genes with zero copy,13genes with single copy,17genes with2copies,14genes with2or more copies.2. Based on the5KB sequences upstream and downstream of102glucosinolate biosynthesis genes in Chinese cabbage,237SSR loci were developed,16of them without SSR,72of them with1~4SSR loci,5of them with5SSR loci,4of them with6SSR loci,4of them with7SSR loci,1of them with8SSR loci.Within developed SSRs, a total of four repeat types were found, in which singlenucleotide repeat was the most, with value of122, accounting for51.4%, thedinucleotide type was the second is, with value of48, accounting for20.3%, trinucleotide repeat types was the least, with value of7, accouting for3.0%of thetotal number of SSR,60of them has no obvious repeat types.3. Within the86genes with SSR loci,77genes could be designed specific SSRprimers. There were49specific primers had good amplyfication in75Chinesecabbage inbred lines.16of them with polymorphism, accounting for20.8%,18ofthem with polymorphism, accounting for23.4%,15of them with addional bands,accounted for19.5%,28of them without amplification products, accounting for36.4%. Finally,11primers were amplified in75Chinese cabbage inbred lines,26alleles were detected, with average2.4alleles of each primer.4. Within developed177InDeL primers,161of them had good amplification in75Chinese cabbage inbred lines, accounting for91.0%of the total primers,61ofthem with clear polymorphism products, accounting for34.5%. In total161polymorphic bands were dtecetd using61pairs of primers, with average2.6bandsof each primer. And95of them were detected only1band,31of them were detected2bands,24of them were detected3bands,6of them were detected4bands.5. In seedling stage, the glucosinolate level in leaves of68Chinese cabbageinbred lines was determinated. Results showed that there were clear differences indifferent genotypes, with total glucosinolate ranging from0.734to11.189μmol/gDW. The contect of Indole family was higher than in aliphatic and aromatic gourp,with proportion of49.9%,39.8%and10.6%, respectively. Among them, GBC hadthe highest content, accounting for27.0%, the contect of4me and GBN contentwas the second, accounting for18.0%and15.9%of the total content,4OH contentwas the lowest, accounting for only0.6%of the total amount.6. In the mature period, the glucosinolate level in leaves of68Chinese cabbageinbred lines was determinated. Results showed that there were clear differences indifferent genotypes, with total glucosinolate ranging from1.918-17.357u mol/gDW. The contect of indole and aliphatic family was higher than in aromatic gourp,with proportion of48.6%,41.7%and9.7%, respectively. Among them, PRO, GBN4me and GBC had compatable content, accounting for19.8%、19.7%、19.6%and19.0%, respectively,4OH content was the lowest, accounting for only0.7%of thetotal amount. The level of total glucosinolate and components in mature stage washigher than in seedling stage, with almost2folds.7. Using GLM association mapping analysis, On the basis of statisticalprobability P value of0.01,97IDENL and SSR molecular markers associated withglucosinolate were detected, which werte distributed in10chromosomes of Chinesecabbage. The most of detected associations were realetd to4OH, NAP and GBN. |
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