基于BSA-seq技术的青花菜花球蜡质基因初定位及鉴定研究

Wax Genes in Broccoli Curd:Primary Localization and Identification Based on BSA-seq Technology

为了发掘调控青花菜花球蜡质合成的相关基因,以青花菜有蜡粉野生型(WT20)为父本,无蜡粉突变体(MT20)为母本构建F2群体,基于F2构建极端有蜡混池和无蜡混池,利用BSA-seq技术对亲本混池和F2极端混池进行混池测序,通过ΔSNP-index算法初步定位与蜡质合成关联的区间,根据基因注释信息预测候选基因,并采用qRT-PCR分析候选基因在亲本中的表达情况。结果表明F2分离群体中有蜡粉株与无蜡粉株分离比为2.87:1,符合单基因隐性核基因遗传规律;两个混池间共获得高质量SNP位点2,045条、InDel位点687条,基于SNP-index关联分析将候选基因定位在4号、7号和8号染色体的3个区间,包含93个基因。进一步对候选区域内的基因进行KEGG、GO、GOC数据库分析发现0.12 Mb和1.07 Mb的区段内92个基因获得注释,对区间内可能跟蜡质合成相关的11个基因进行定量分析,筛选出4个差异显著表达基因BoAPC3、BoCDH1、BoEEF1A和BoAHP4。本研究结果可以为进一步研究青花菜蜡质合成通路提供新思路,以及为选育高品质青花菜奠定基础。

In order to explore the related genes regulating the synthesis of wax in broccoli flower bulbs,the F2 population was constructed using the broccoli waxy wild type(WT20)as the male parent and the waxless mutant(MT20)as the female parent.Based on F2,extreme wax-pool and no-wax-pool were constructed.The parent pools and F2 extreme pools were sequenced by BSA-seq technology.The intervals associated with wax synthesis were initially located by theΔSNP-index algorithm.Candidate genes were predicted based on genes annotation information,and the expression of the candidate genes in the parents was analyzed by qRT-PCR.The results showed that the segregation ratio of wax-powdered plants to no-wax-powdered plants in the F2 segregation population was 2.87:1,which conformed to the genetic law of single-gene recessive nuclear genes.A total of 2,045 high-quality SNP loci and 687 InDel loci were obtained between the two pools.Based on SNP-index association analysis,the candidate genes were located in three intervals on chromosomes 4,7 and 8,including 93 genes.Further analysis of genes in the candidate regions in KEGG,GO,and GOC databases revealed that 92 genes in the 0.12 Mb and 1.07 Mb segments were annotated.Quantitative analysis was performed on 11 genes that might be related to wax synthesis in the intervals,and 4 differentially significantly expressed genes,BoAPC3,BoCDH1,BoEEF1A,and BoAHP4,were screened out.The results of this study can provide new ideas for further research on the wax synthesis pathway of broccoli,and lay a foundation for the breeding of high-quality broccoli.