番茄果实硬度重要调控基因的克隆与功能分析

Tomato fruit firmness, one of the most important agronomic trait, highly correlated with the thickness, compression resistance and puncture resistance of pericarp. Several quantitative trait loci (QTLs) related to tomato fruit firmness have been identified but the molecular markers derived from QTL analysis are not available in marker-assisted selection (MAS). Identification of molecular markers more tightly linked to the fruit firmness is necessary. Previously, a cross was made between tomato cultivar 'Moneymaker' and its wild relatives Solanum.lycopersicum var. cerasiforme accession LA1310 to develop a recombinant inbred lines (RILs). Based on the phenotype of fruit firmness and high-throughput DNA sequencing for the subset of 235 individuals in RILs, a major QTL was detected using genome-wide association study (GWAS). In the following study, we plan to fine map and clone key genes within major QTL associated with fruit firmness using SNP and Indel markers in mapping population. Furthermore, the map-based cloning of the candidate genes will be validated by transgenic gene editing techniques and complementation experiments. These achievement will contribute to the marker-assisted selection in high firmness tomato.

果实硬度是评价番茄品质优劣的重要性状之一，该性状与果皮厚度、耐压力及耐穿刺力等指标显著相关。国内外通过果实硬度相关性状的遗传分析，初步定位了一些控制番茄果实硬度的QTL位点，但是仍缺乏可应用于分子育种的分子标记，因此克隆控制番茄果实硬度的重要调控基因对加快高品质品种的选育具有重要的意义。本研究在前期以番茄栽培种Moneymaker和野生种LA1310为亲本材料，构建包含235个株系的重组自交系RILs群体，通过对RILs群体的重测序及果实硬度相关表型的全基因组关联分析，鉴定了调控果实硬度的重要QTLs。本项目计划通过构建定位群体，利用在主效QTL区间内特异开发SNP或InDels标记，精细定位和克隆调控番茄果实硬度的关键基因；同时，通过基因编辑系统及互补实验对候选基因进行功能验证，从而解析其在调控果实硬度中的作用，为番茄果实品质育种提供理论指导，加快高硬度番茄品种的选育。

果实硬度是决定番茄品质的重要性状之一，直接影响番茄的经济价值。本项目以重组自交系群体为材料，利用全基因组关联分析结合遗传分析方法，精细定位并克隆了一个调控果实硬度的关键基因，Fir10.1。为验证其功能，我们利用CRISPR技术获得两个独立的突变体株系，分别命名为fircr-1/-2。与野生型相比，突变体的果实硬度显著增加，说明该基因确实是调控果实硬度的关键基因，且其缺失能够使果实硬度增加。通过对近等基因系果皮进行细胞学和代谢组学分析发现，具有MM基因型株系是通过提高角质和蜡质含量来增加果皮角质层厚度，从而使果实硬度增加。综上所述，我们的项目为深入了解果实硬度的生物学机制提供了新的见解，也为提高番茄果实硬度提供了新的策略。