Mapping Of The Early Bolting Mutant Gene Ebm9 In Chinese Cabbage And Analysis Of Thecandidate Gene

Timely bolting is crucial to the production and breeding of Chinese cabbage（Brassica rapa ssp.pekinensis）.Creating a variety of mutants related to bolting,cloning mutant genes,and analyzing the reasons for the changes in bolting traits of Chinese cabbage caused by their mutations are important ways to study the molecular regulatory mechanism of Chinese cabbage bolting and flowering.In this study,a stable genetic early bolting mutant of Chinese cabbage was obtained using EMS mutagenesis technology,and then using Mut Map technology to locate the mutant gene,clarify its expression pattern,and identify the function of the candidate gene BrPMEI51.The relevant results are as follows:1.Compared with the wild type‘FT’,the early bolting mutant has no obvious balling phenomenon during autumn cultivation and bolts directly.Genetic analysis showed that the early bolting trait of this mutant was controlled by a single recessive nuclear gene,named ebm9（early bolting mutation gene 9）.2.The F₂segregating population was constructed using wild-type and mutants,and early bolting single plants in the segregated population were identified to construct mixed pools.The mutants,wild-type and mixed pools were re-sequenced respectively,and the mutation sites were located with the help of Mut Map strategy Finally,the mutation site was located at 1532588bp of chromosome A02,which was located on the promoter of the pectin methylesterase inhibitor gene BrPMEI51（Bra A02g003140.3C）,and a single-base mutation（C→T）occurred.3.The qRT-PCR analysis showed that the expression level of BrPMEI51 gene on mutant ebm9 and wild type’FT’was similar,but the expression level of flower on mutant ebm9 was higher than that of wild type;GUS histochemical staining method indicated that genes are expressed in roots,inflorescences,and leaves;The subcellular localization results show that BrPMEI51 protein is distributed in the cytoplasm,and the specific organelles in which it is located need further study.4.The BrPMEI51 overexpression vector was constructed and transformed into Colombian ecotype（col-0）Arabidopsis.The results showed that compared with the wild type,the bolting of the Arabidopsis plants overexpressing BrPMEI51 was significantly earlier.Therefore,it is speculated that the function of the BrPMEI51gene is closely related to the bolting traits of the plant.