小粒野生稻抗白背飞虱新基因WBPH9的克隆与抗虫机理分析

Whitebacked planthopper (WBPH) is an important pest for rice production, not only harms rice plan directly, but also intermediates other pathogen or virus during feeding process, and causes serious threat to the safe production of rice in China. Identification of new resistance gene resources and breeding for resistant varieties are the most economic and effective control means. In the previous work, we detected a novel gene WBPH9 for resistance to WBPH from K1561, an introgression line of Oryza minuta, which has been fine mapped to 60 kb region on Chromosome 6 and obtained the candidated gene. In this project, firstly, we are planing to conduct functional complementarity analysis, spatiotemporal expressing analysis, and subcellular localization analysis for WBPH9. The objective of these works are to invest the biological function and effective tissues of WBPH9. Secondly, we will analysis the host selection, feeding behavior, food intake, survival rate, population growth rate and spawning force of WBPH in transgenic plants and near-isogenic line with WBPH9, in order to resolve the resistance mechanism of WBPH9 against WBPH. Finally, we will evaluate and verify breeding value of WBPH9 in rice transgenic breeding. The fulfillment of the project will promote the understanding of insect resistance mechanism in rice in theory. In practice, WBPH9 can also be applied to molecular breeding for resistance to WBPH as a novel gene resource.

白背飞虱是水稻生产上一种重要的害虫，不仅直接危害水稻，还在取食过程中传播其他病害和病毒，严重威胁到我国水稻的安全生产。发掘和鉴定新的抗虫基因资源，选育抗虫品种是最经济有效的防治手段。在前期研究中，我们从小粒野生稻渗入系K1561中发现了一个新的抗白背飞虱基因WBPH9，将其定位在第6号染色体上60kb区间内，通过基因序列比对确定了其候选基因。本项目拟在此工作基础上，进一步开展基因功能互补分析、基因时空表达分析和亚细胞定位分析，明确WBPH9的生物学功能和作用部位；并从宿主选择性、取食行为、取食量、存活率、群体增长率和产卵力入手，确定WBPH9防御白背飞虱的抗性机理；同时通过转基因水稻和近等基因系对白背飞虱抗性鉴定，进行WBPH9转基因育种价值评价与验证。本项目的完成，在理论上可以增进对水稻抗虫机制的理解；在应用上可以为水稻抗白背飞虱的分子育种提供基因资源。

白背飞虱是水稻生产上一种重要的害虫，不仅直接危害水稻，还在取食过程中传播其他病害和病毒，严重威胁到我国水稻的安全生产。发掘和鉴定新的抗虫基因资源，选育抗虫品种是最经济有效的防治手段。本项目在前期工作基础上，首先，构建了WBPH9 基因功能互补载体和 CRISPR/Cas9基因编辑载体，获得了转基因植株，通过对转基因T0、T1和T2代进行表型和基因型鉴定与分析，完成了WBPH9 的功能验证；通过GUS报告基因系统检测WBPH9在K1561和G1025不同组织中的时空表达，同时，采用荧光定量PCR检测了WBPH9在不同组织器官中的表达水平；其次，利用WBPH9近等基因系和转基因植株为鉴定材料，通过对白背飞虱若虫、成虫和产卵的忌避性，以及蜜露量、生长速率、产卵量、孵化率等抗生性指标的测定分析，明确了WBPH9对白背飞虱的抗性机理兼具忌避性和抗生性；最后，通过对WBPH9 转育水稻材料的白背飞虱抗性鉴定，明确了WBPH9 在不影响水稻生长发育特性和杂种优势表现的前提下表现出对白背飞虱的抗虫效果，同时确认其在抗白背飞虱水稻育种中具有重要的应用价值。本项目的完成，在理论上可以增进对水稻抗虫机制的理解；在应用上可以为水稻抗白背飞虱的分子育种提供基因资源。