水稻抗褐飞虱基因BPH9显性上位基因DE9的克隆及其调控机理研究

Rice (Oryza sativa L.), an important cereal crop in Asian-Pacific region, is a stable food resource for more than half the population of the world. The brown planthopper (BPH) has become the most destructive and a serious threat to the rice production in Asia. Breeding the resistant varieties with improved host resistance is the most effective and ecosystem-friendly strategy of BPH biological management. To date, at 31 BPH-resistant genes have been identified in the cultivated rice and wild Oryza species, fourteen of these resistance genes have been cloned by map-based cloning, which provide resistance genes for marker assisted selection (MAS) breeding. Several studies showed most of the donors showing durable resistance over the years and biotypes carry many minor QTLs in addition to one or more major genes. Hence identification and use of the QTLs along with major genes will help in developing durable resistance. Using different mapping populations from crosses of susceptible and resistant varieties, more QTLs were detected on all rice chromosomes except 5 and 9. However, there are no further reports of those minor QTLs, possibly due to the complex inheritance of the BPH resistance. .In our previous study, we introgressed BPH9 into indica 9311 and japonica Nipponbare backgrounds using marker-assisted selection (MAS). The resulting near isogenic lines in 9311 and Nipponbare backgrounds were named as NIL-BPH9-9311 and NIL-BPH9-Nip, respectively. NIL-BPH9-9311 showed high levels of resistance to BPH. However, NIL-BPH9-Nip was susceptible to BPH. Significantly, the hybrids derived from NIL-BPH9-Nip and 9311 also conferred high level of resistance to BPH, suggesting a dominant epistatic locus from 9311 contribute to resistance of BPH9. This locus was designated as DE9 (Dominant Epistatic gene of BPH9). We used map-based cloning strategy to isolate DE9. We have performed high-resolution mapping to narrow DE9 to the 20kb genomic region, which contains only one predicted gene. Complementation tests with this candidate gene for DE9 are undergoing. In this proposal, we will focus on identification and characterization of DE9. We will further elucidate the molecular mechanisms of DE9 in regulating the resistance of BPH9 against BPH and facilitate the development of durable resistant varieties for sustaining control of BPH pest in rice crop.

水稻是我国最重要的粮食作物，褐飞虱是水稻生产的首要害虫。遏制褐飞虱的发展和危害是保障我国乃至亚洲水稻生产安全的重大需求。从稻种资源中发掘、定位和克隆的抗褐飞虱基因已开始应用于抗褐飞虱育种实践。以往研究发现遗传背景或QTL位点能够显著影响主效抗褐飞虱基因的抗性水平和持久性，对培育稳定持久抗虫品种同样至关重要。但至今为止并未见到这些位点的后续研究报道。我们在前期研究中发现籼稻9311中促进BPH9抗褐飞虱功能发挥的有利显性上位位点DE9，并将其精细定位在单个候选基因区段。本项申请拟在此基础上进一步克隆DE9基因，解析DE9基因的生物学功能，进行DE9基因的分布、起源与进化研究。明确DE9与BPH9之间的互作及调控关系，阐明DE9调控BPH9抗褐飞虱功能的分子机理。本项目的研究结果将为BPH9在抗褐飞虱育种中的应用提供理论依据，也将为利用主效抗褐飞虱基因培育持久稳定抗虫品种提供重要的参考和借鉴。

水稻是我国最重要的粮食作物，褐飞虱是水稻生产的首要害虫。遏制褐飞虱的发展和危害是保障我国乃至亚洲水稻生产安全的重大需求。从稻种资源中发掘、定位和克隆的抗褐飞虱基因已开始应用于抗褐飞虱育种实践。以往研究发现遗传背景或QTL位点能够显著影响主效抗褐飞虱基因的抗性水平和持久性，对培育稳定持久抗虫品种同样至关重要。但至今为止并未见到这些位点的后续研究报道。项目在前期研究中发现籼稻9311中促进BPH9抗褐飞虱功能发挥的有利显性上位位点DE9，并将其精细定位在单个候选基因区段。本研究进一步应用图位克隆策略克隆了DE9，发现DE9是Karrikin信号通路重要基因OsSMAX1，应用酵母双杂交及水稻原生质体亚细胞定位实验明确了DE9与TPR2与D14L的互作关系，且Karrikin信号通路关键基因D14L在蛋白水平上受褐飞虱诱导表达，表明Karrikin信号通路参与调控抗虫性。本研究证实DE9基因除影响BPH9抗褐飞虱功能外，还影响农艺性状表型，表明DE9可能在植物生长发育与抗性间的平衡起作用。BPH9定位于内膜系统，DE9定位于细胞核，二者的定位模式不同，并不直接互作。本研究筛选了DE9基因的互作蛋白、结合高通量转录组分析DE9调控的基因网络和信号通路，表明DE9作为转录抑制因子调控下游目标基因的表达进而调控BPH9的抗褐飞虱功能。