| Brassica rapa, comprising many important vegetable and oil crops, has a long cultivation history and extensive plant areas in China. With the improvement of people’s living standard, the nutritional quality traits of Brassica rapa have been paid more and more attention. Glucosinolate, an important secondary metabolite of Brassica rapa, plays essential roles in growth and development as well as provides indispensable resources for flavor and health care. In particular, sulforaphanethe, degradation products of glucoraphanin, can inhibit the development of various cancers such as bladder cancer,lung cancer and gastric cancer. However, trace amounts of or no glucoraphanin were detected in Brassica rapa. Therefore, it is necessary to understand the genetic diversity of glucosinolates and the biological synthesis pathway of glucosinolates in Brassica rapa,which lays a foundation for mining of beneficial glucosinolates and key regulatory genes and provides a theoretical basis for the cultivation of high beneficial glucosinolate. In the current study, the UPLC-qTOF-MS method was used to analyze the diversity of glucosinolates and the difference of glucosinolate in different subgroup of Brassica rapa. Furthermore, GWAS was conducted to identify the candidate genes for glucosinolates. Finally, molecular marker assisted breeding was used to improve the content of glucoraphanin in Brassica rapa. The main results of this study are as follows:1. Using UPLC-qTOF-MS,a total of 32 different glucosinolates which included 24 aliphatic glucosinolates, five indolic glucosinolates and four aromatic glucosinolates were detected in the collection of 99 Brassica rapa. Among these glucosinolates, 18 kinds of glucosinolates were newly identified in Brassica rapa. Seven major glucosinolates (NAP, GBN, GST, NEO, I3M, GRA and PRO)were used to analyze the difference of glucosinolate in seven subspecies of Brassica rapa. The high content of indolic glucosinolate NEO and beneficial indolic glucosinolate I3M was found in Chinese cabbage; the high concentration of beneficial aliphatic glucosinolate GRA and low concentration of detrimental glucosinolate PRO was detected in Yellow Sarson.2. The mGWAS study was conducted using 99 Brassica rapa accessions and 32 kinds of glucosinolates. A major polymorphic loci (InDel) of Bra016433 was associated with 4OH-I3M and 4MO-I3M glucosinolate in Brassica rapa. Furthermore, we identified other 21 locis for various glucosinolates. Importantly,a stop mutation of BrAOP2.2(Bra000848) was detected which was significantly associated with the beneficial glucosinolate GRA.3. AOP2 gene was a key gene which was associated with the accumulation of beneficial glucosinolate GRA. The beneficial glucosinolate GRA was highest in Yellow sarson accession through analyzing the content of glucosinolates in different subgroups of Brassica rapa. In addition,we investigated sequence variations of all threeBrAOP2 genes between Yellow sarson type ’R-O-18’and ’Chiifu-401/42’. The sequence variation of BrAOP2.2 and BrAOP2.3 in ’R-O-18’ resulted in the premature translation termination products, and the enzymatic activity showed that BrAOP2.2 and BrAOP2.3 protein abolished catalysis activity. Finally, the replacement or introgression of single non-functional braop2.2 and braop2.3 locus led to the enrichment of glucoraphanin in ’L58’ through marker-assisted backcrossing. |
