Microarray-assisted identification of genes associated with blast resistance in rice

The authors are interested in determining the genetic control of durable resistance against rice blast disease. Previous work showed that SHZ-2, an indica cultivar grown in China, has broad spectrum resistance to multiple races of the blast pathogen. A number of QTL has been mapped using recombinant inbred lines derived from SHZ-2. The authors crossed SHZ-2 to TXZ-13, a blast susceptible but high-quality variety to produce two BC3 lines, BC10 and BC116. These two lines showed strong to moderate blast resistance over eight cropping seasons in the field. In particular, BC10 has been successfully used in the hybrid rice program in Guangdong Province to produce high-yielding and blast resistance hybrids. To further dissect the QTL responsible for durable blast resistance, 451 BC4F3 lines were developed by backcrossing BC10 and BC116 to TXZ-13. The BC4 F3 population consisted of 244 lines from BC10 and 207 lines from BC116. The two BC4 populations were evaluated for blast resistance in the greenhouse and the blast nursery at IRRI, Philippines. The chromosomal introgression from SHZ-2 was identified by using a genome-wide, genotyping microarray produced at University of Arizona. This array consists of 880 oligos that detect single feature polymorphisms (insertions and deletions) in unique genes evenly spaced along the chromosomes (medium spacing about 250 kb). Genome DNA of BC10 and BC116 were enzyme-digested and labeled with Cy5; TXZ-13 was digested and labeled with Cy3. Hybridization was done by pairing labeled DNA of BC10 and BC116 with that of the recurrent parent TXZ-13. Graphical genotypes at BC10 and BC116 were produced based on hybridization patterns using the software of Graphical Genotyping (GGT) version 2.0 (http://www.dpw.wau.hl/pv/pub/ggt/www.plantbreeding.nl). Based on data from replicated hybridization experiments (4 times), the authors detected 20 and 17 regions of SHZ-2 introgression segments in BC10 and BC116, respectively. Some of the regions of SHZ-2 introgression shown by microarray data were checked by SSS markers located near the introgression regions. Of 27 SSR markers tested, 10 were consistent with the microarray data; the other SSR markers were monomorphic and hence not informative. To determine the relationship between introgression regions and blast resistance, extreme resistant and susceptible BC4F3 lines were selected for analysis. Based on genome-wide graphical genotypes and gene expression data of SHZ-2, the authors selected a panel of genes to monitor the introgression of SHZ-2 alleles in BC4F3 lines by TILLING (a procedure that detects SNP between two target sequences). Through a combination of genome-wide genotyping and detection of specific alleles, the authors expect to be able to determine the genetic regions or candidate genes responsible for blast resistance in the advanced backcross.