Identification Of The Quantitative Trait Gene (QTG)with Resistance To Sheath Blight In Rice

Sheath blight (SB) is one of the three most serious diseases in rice worldwide, causing huge yield losses and bad quality when it developed severely. Resistance to rice sheath blight is a typical quantitative trait. To date, no varieties were found of high-resistance or immunity to sheath blight disease fungus. Therefore, mining quantitative trait locus QTL) in some medium resistant rice varieties is of important practical significance, and is also advantage of investigating the mechanism of quantitative resistance.In our previous study, we had mapped the QTLqSB-11Le into a74kb region on chromosome11of Lemont, Through repeat inoculation test for some recombinant chromosome segment substitution lines (CSSLs), we further confirmed the accuracy of the above mapping region. which contains11predicted ORFs (Open Reading Frame). Five of them have annotations. Semi-quantitative RT-PCR was used to analysis the expression pattern of some ORFs at different time-intervals after fungus inoculation. Results indicated that all five ORFs with annotated information are expressed genes, other ORFs don’t express. By sequencing (include coding regions and2kb upstream and lkb downstream of each gene), we found that the sequence of ORF3was same in parents, while the others were different in parents and were considered as the candidates of qSB-11LeSpecific PCR primers located in coding region were designed for amplifying specific region of each of four candidates. Using Lemont DNA (containing qSB-11Le) as template, four specific PCR products, were obtained and introduced into RNAi vector. Also, the over-expression constructs and YFP.(yellow fluorescent protein)-fused expression constructs for ORF10and ORF11were developed. Through agrobacteria-mediated transformation, the constructs were introduced into rice cultivar Lemont. For RNAi constructs of ORF1, ORF6and ORF10and overexpression construct of ORF10, we had obtained some positive transgenic plants. All others constructs were being transformed. By using gene gun transformation method, the YFP-fused expression constructs were introduced into onion cells, and the transient expression indicated that ORF10protein was located in the nucleus as well as the cell membrane.Some of RNAi and overexpression transgenic plants were inoculated SB fungus. The Lemont and its isogenic line NIL-qSB-11Tq (carrying qSB-11Le susceptibility alleles) were used as relatively resistant and susceptible controls, respectively. The disease lesion length and disease rating of each plant were measured/rated at7days after inoculation and30days after heading, respectively.Results showed that, no apparent difference was found between RNAi transgenic plants of ORF1and ORF6and Lemont; whereas, RNAi plants of ORF10presented more susceptible than Lemotn. Combined with the previous studies, we preliminary think that the ORF10is the most likely qSB-11Le candidate gene.Generally, large population and multi-replicates were required when identifying the phenotype of quantitative resistance. In this studies, the experimental populations for each transgenic plants were relatively small, and over-expression transgenic plants were not be inoculated, so the results should be verified in future studies. Meanwhile, ORF10complementary vector being constructed, complementary transformation is underway.The present study has laid a solid foundation for these works, which will accelerate the qSB-11Le characterization.