Isolation and identification of a gene in response to rice blast disease in rice

We combined cDNA amplified fragment length polymorphism (cDNA-AFLP) with bulked segregant analysis (BSA) to detect genes that control rice blast (Magnaporthe grisea) resistance in a double-haploid (DH) population derived from a cross between a blast-resistant variety, Zhai Ye Qing8 (ZYQ8), and a blast-susceptible variety, Jin Xi17 (JX17). In cDNA-AFLP analysis between a blast resistance (R) pool and a blast susceptibility (S) pool from the DH population, 12 transcript-derived fragments (TDFs) that were present in only one of the two pools were detected, 8 of which were from the R pool and 4 from the S pool. Mapping analysis of these TDFs by using the DH mapping population showed that five of them, R1, R8, S9, S16 and S17, were located on chromosome 1. Sequence comparison and allelic analysis showed thatR1/S16 and R8/S9 were two pairs of allelic genes. The full-length cDNA sequences of R1/S16, S17 and R8/S were obtained through cDNA library screening, in which only the expression level of R8 cDNA was up-regulated by inoculation with the blast isolate zh10814 and not affected by mock treatment, suggesting that R8 was implicated in the signaling pathways of the rice blast resistance reaction. Protein function prediction showed that R8 cDNA encodes a protein with high identity to a putative calmodulin-binding protein in Arabidopsis thaliana which belongs to the P-loop-containing nucleotide triphosphate hydrolases superfamily that contains a number of various kinases.