Gene Mapping and Blast Resistance Analysis of a Lesion Mimic Mutant spl52 in Rice (Oryza sativa)
LI Yuan-Yuan1, LI Xing1, HUANG Qing-Xiong2, QI Pan1, YIN Wu-Zhong1, ZHANG Jie1, CHEN Hong1, YANG Guo-Tao1, PENG You-Lin2,*, HU Yun-Gao1,*
1 Rice Research Institute of Southwest University of Science and Technology, Mianyang 621000, China; 2 Yazhou Bay National Laboratory, Sanya 572024, China
Abstract：Rice (Oryza sativa) blast is one of the factors affecting the stable yield of rice. The lesion mimic mutant is an ideal material for studying the molecular mechanism of crop disease resistance. The analysis of the disease resistance gene of the mutant is the basis for cultivating stable and disease-resistant rice. In this study, a stable genetic lesion mimic mutant spl52 (spotted leaf 52) was obtained by ethylmethylsulfone (EMS) mutagenesis of japonica rice 'Wuyunjing 21'. The effective panicle number, grain number per panicle, 1000-grain weight and seed setting rate of the mutant plants at maturity stage were significantly lower than those of the wild type. From the early tillering stage, the leaf tip of mutant spl52 appeared disease spots. By 3,3'-Diaminobenzidine (DAB) staining, enzyme activity assay and 2',7'-dichlorodihydrofluorescein diacetate (H2DCFDA) detection results showed that the mutant spl52 lesion leaves showed reactive oxygen species accumulation, and programmed cell death activation was found by TUNEL assay. Transmission electron microscopy analysis of wild-type and mutant leaves showed that the mutant chloroplasts were smaller oval, and the mitochondria were hollowed out. Genetic analysis showed that the phenotype of the mutant was controlled by a pair of recessive nuclear genes. Six possible candidate intervals were found by Mutmap analysis, and the candidate genes were located between RM3.5-RM4.3 on chromosome 6 by map-based cloning. The candidate genes were analyzed, and no reported lesion-like and rice resistance-related genes were found. The leaves of wild-type and mutant plants were inoculated with rice blast fungus, and the results showed that the mutant plants were more sensitive to rice blast fungus (Magnaporthe oryzae). The results of qRT-PCR showed that the expression levels of OsPR1a, OsPR1b, JIOsPR10, OsWRKY6 and OsRSR1 which positively regulated rice resistance in the mutant were significantly lower than those in the wild type. The results showed that the mutant gene was a new gene that had not been reported and was involved in the regulation of rice blast.
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