兰麦抗麦长管蚜基因SaLMZS的精细定位及候选基因分析

English grain aphid (EGA) (Sitobion avenae F.) is one of the most important pests in wheat production. Wheat landrace of Shaanxi Province Lanmai (Shaanxi Zhashui), showed highly resistant to EGA both in seedling and adult stage, is an ideal material to dig EGA resistance genes. The RNA resistant bulk (R-bulk) and susceptible bulk (S-bulk) were constructed for BSR-Seq in the population of F2:3 NIL-S/NIL-R derived from the cross Xiaoyan 6/Lanmai (ShaanxiZhashui). In addition, a pair super DNA “R-bulk vs S-bulk” were performed from the population of F2:3 from the cross Polan305/Lanmai (ShaanxiZhashui) for SLAF-seq analysis. New EST-STS and SNP primers were developed with genome comparative analysis method and RNA-Seq and SLAF-seq sequencing technique.The precise genetic linkage map of SaLMZS gene was drawn with those primers above which are newly desighed. Characterize the collinearity of the SaLMZS region in wheat genome and those in the genomes of model crop. Identify candidate genes of SaLMZS in the target region, and clone the candidate genes in common wheat. Through realtime PCR (QRT-PCR) and virus induced genetic silence (VIGS) method, the gene expression characteristics and functions of candidate genes induced by EGA pathogen were investigated. This project is expected to obtain one or two wheat EGA-resistance candidate genes, to clarify their mechanism in the process of resistance to EGA. This work will enrich genetic resources of wheat EGA-resistance, provide references for researching resistance mechanism of wheat EGA and carry out anti-aphid molecular breeding.

麦长管蚜（Sitobion avenae）是我国小麦生产上主要害虫之一，地方小麦品种兰麦（陕西柞水）在苗期和成株期均高抗麦长管蚜，是挖掘麦长管蚜抗性基因的理想材料。本项目以兰麦（陕西柞水）与感蚜材料波兰305杂交，以及其近等基因系N09H21SA和N09H21sa杂交分别构建遗传分离群体，基于集群分离分析（BSA）策略，运用BSR-Seq和SLAF-seq-BSA混池测序技术，进行SNP引物的开发和标记筛选，绘制SaLMZS基因的精细连锁遗传图。利用小麦基因组信息对目标区域内进行候选基因预测，确定并克隆SaLMZS的候选基因。利用实时荧光定量PCR与病毒诱导基因沉默(VIGS)的技术，对己克隆的候选基因进行了基因表达特性与抗蚜功能分析。本项目预期获得SaLMZS的高密度遗传图谱和1-2个小麦抗麦长管蚜候选基因，并阐明候选基因在小麦抗麦长管蚜过程中的作用机理。

麦长管蚜是我国小麦上最重要的害虫，严重威胁我国小麦生产与粮食安全。地方小麦品种兰麦在苗期和成株期均高抗麦长管蚜，是良好的抗源。本研究结果显示，其对麦长管蚜表现出较强的排趋性和抗生性，但是耐受性较差。基于麦长管蚜抗性鉴定结果进行遗传分析，表明兰麦的麦长管蚜抗性由1对显性单基因控制（暂命名为Sa3）；通过BSA结合BSR-Seq和芯片测序并利用KASP技术，获得该抗蚜基因的高密度连锁图谱，将兰麦抗蚜基因Sa3定位于1A染色体上AX-109079516与AX-111118840之间。将两标记区间内的差异SNP比对到小麦基因上，经基因注释和表达分析，确定TraesCS1A01G253200（Protein phosphatase 2C family）和TraesCS1A03G0124900（lipoxygenase）为兰麦抗蚜候选基因。并对抗蚜候选基因TtPP2C和TtLOX进行了克隆及功能验证。本研究为解析农作物抗蚜机理及开展抗蚜分子育种奠定基础，对提高小麦抗蚜育种具有重要的理论和实践意义。