Fine Mapping And Candidate Gene Analysis Of Major Margin Cracking QTL Lob10.1 In Chinese Cabbage

Leaf-lobed is an important agronomic trait,which is related with photosynthesis and plant stress resistance in crops.In order to fine map and isolate the lobed-leaf gene in Brassica rapa,an NIL-F2 population derived from the cross between the recurrent parent Rc Br and lobed-leaf main effect QTL near iso-genic lines（NIL）were constructed.As a Result,one major QTL was identified on A10,and two SNP markers were developed based on RNA-seq data.In light of annotations of the genes in candidate region,a leaf morphological development related gene Bra009510,was primary identified as the candidate gene.The main Results are as follows:1.Fine mapping of major QTLThe NIL-F2 population was used as the material,and the dominant plants were used to verify the two labeled primers of WXIn Del-17 and WXIn Del-76.The recombinant plants were screened and verified by progeny.The genotypes of the whole plant were also verified.Combined with the F2 generation single genotype and the progeny leaf shape separation,the gene was finally positioned between Br ID10909 and WXIn Del-48 with a physical distance of139.7 kb.2.RNA-seq data analysisIn this study,a transcriptome library of NIL and P2 was constructed using Brassica rapa V₂.5 as a reference genome.A total of 124 differentially expressed genes were detected.Analysis of its function revealed no differentially expressed genes associated with leaf edge cleavage,presumably because the sampling period was too late.Further analysis of the SNP loci between the parents,a total of 1017 homozygous differential SNP loci were detected,of which 880 were located in A10,91 of which were located in the localization interval.Based on the data,two pairs of polymorphisms were developed.SNP markers for further fine positioning later.3.Screening and prediction of candidate genesThere are 25 candidate genes in this localization interval.The Bra009510 gene is similar to the Arabidopsis AT5G03790 gene in the localization interval.The previous studies showed that AT5G03790 can make the Arabidopsis leaf margins jagged.It is nicked and can inhibit the formation of leaves and temporal lobe.As a popular gene for leaf margin cleavage,Bra009510 gene has been localized by many people,but its regulation mechanism of leaf cleavage has not been resolved.4.Cloning and identification of candidate genesBra009510 is a popular gene for leaf cleavage traits.Primers designed for Bra009510.By cloning and sequencing analysis,it was found that P2 has a base insertion in the exon region of Bra009510 gene,which may Result in a frameshift mutation.Therefore,it is inferred that the gene is large.Candidate gene for B.rapa leaf lobes.