砂梨抗黑斑病基因精细定位及其候选基因功能初步解析

Pear black spot disease, caused by Alternaria alternata is one of the most serious diseases in sand pear. Molecular breeding of disease-resistant varieties has been the most effective and economic way to control the disease. In order to obtain the sand pear disease resistant related genes, and analysis preliminary function of resistance to the black spot, in this study, "Jinjing Pear" which is high resistance cultivar to the black spot, "Hongfen pear" which is high susceptibility cultivar to the black spot, and their hybrid F1 as experimental materials, resistance associated area to black spot disease will be screened by super BSA analysis of hybrid F1 generation using specific locus amplified fragment sequencing. Candidate genes will be identified and fine mapping combined with the analysis of previous transcriptome analysis and super BSA. Analysis of tissue specific expression of candidate genes in different tissues and organs of pear and analysis of the induced expression patterns of black spot at different time periods were studyed by qRT-PCR, and the primary function of disease resistance was verified by Transformation of Arabidopsis thaliana. The research lays the theoretical foundation for study on disease resistant mechanism of sand pear and cultivate new resistance varieties to the black spot.

梨黑斑病是危害梨树生长的主要病害之一，植物抗病分子育种是一种经济有效的防治病害的手段。为了获得砂梨抗黑斑病密切相关的基因，并初步解析其抗病功能，本研究以高抗黑斑病梨品种“金晶梨”和高感黑斑病梨品种“红粉梨”及其杂交F1代为实验材料，利用SLAF-seq测序技术对 “金晶梨”和 “红粉梨”杂交F1代的抗病池和感病池进行Super BSA分析，精细定位砂梨抗黑斑病相关联的区域；结合前期转录组获得的抗黑斑病紧密相关基因，确定砂梨抗黑斑病的候选基因；利用荧光定量PCR技术分析候选基因的组织特异性表达和不同时段病原菌诱导表达模式，同时通过转化拟南芥初步验证抗病候选基因的功能。本研究为解析砂梨抗黑斑病分子机理和培育抗黑斑病新品种奠定理论基础。。

梨黑斑病是危害梨树生长的主要病害之一，植物抗病分子育种是一种经济有效的防治病害的手段。本研究以高抗黑斑病梨品种“金晶梨”和高感黑斑病梨品种“红粉梨”及其杂交F1代为实验材料，利用SLAF-seq测序技术对 “金晶梨”和 “红粉梨”杂交F1代的抗病池和感病池进行Super BSA分析，筛选砂梨抗黑斑病相关联的两个区域scaffold 87.0和scaffold 688.0；结合转录组和Super BSA分析进行生物信息学分析筛选出砂梨抗黑斑病候选基因Pbr040066和Pbr037418；利用荧光定量PCR技术分析候选基因Pbr040066和Pbr037418在叶片、一年生枝韧皮部和果皮等3种器官中的相对表达量和在接种0天、1天、2天、3天和4天不同时间段的表达量变化；同时对Pbr040066和Pbr037418基因进行超表达载体构建并转化拟南芥，通过对转化基因拟南芥的叶片接种的表型和基因型分析，初步确定Pbr040066和Pbr037418基因与砂梨抗黑斑病紧密相关。.本研究基于SLAF-seq测序技术利用Super BSA方法，结合前期转录组测序结果筛选砂梨抗黑斑病候选基因。这两种研究技术结合可以在砂梨转录组水平和基因组水平同时进行检测，能更准确筛选砂梨抗黑斑病候选基因，保证结果的可靠性。本研究为解析砂梨抗黑斑病分子机理和培育抗黑斑病新品种奠定理论基础。