Isolation and characterization of genes related to sheath blight resistance via the tagging of mutants in rice

Abstract

Sheath blight disease (ShB) in rice is caused by Rhizoctonia solani. However, there have been no major genes identified from wild rice relatives and cultivated rice that encode resistance to R. solani. In this study, 300 Ds tagging mutants were generated via tissue culture regeneration methods from a Ds (dissociation) insertion line, and R. solani AG1-IA was inoculated to isolate the resistant mutants. The starter line that was homozygous (Ds-s) with a single Ds insertion in chromosome 1 exhibited a similar response to ShB as the non-transgenic plants. A Southern blot analysis showed that the Ds element was actively transposed in 300 R1 lines generated from Ds-s, and R2 plants were generated by selfing using 300 R1 lines with Ds insertions. Out of 300 R2 lines, two lines displayed susceptibility, and two lines exhibited strong resistance to ShB. In addition, 23 mutant lines displayed resistant symptoms, and the remaining 273 lines showed no differences compared to the ShB-susceptible wild-type control plants. Further examination revealed that the Ds gene was inserted at the second intron of either the PHYTOCHROME B (PHYB) or LAZY1 gene in two strong resistant mutants. We showed that the ethylene signaling genes (EIL1, EIL2, EIN2, and ERF63), jasmonic acid synthesis gene (LOX2), and salicylic acid signaling gene PBZ1 were up-regulated in the phyB mutants compared with the wild-type control that is susceptible to ShB. Further genetic study using EIL1 RNAi and EIL1 overexpression plants revealed that EIL1 positively regulates the resistance of rice to ShB. Overall, our analyses provided a basis for the identification of ShB resistance genes and the realization that PHYB negatively regulates the resistance of rice to ShB via the suppression of ethylene signaling.