Fine Mapping And Candidate Gene Identification At The Xa14 Locus For Bacterial Blight Resistance In Rice (Oryza Sativa L.)

Bacterial blight, caused by Xanthomonas oryzae pv. Oryzae, is one of the most serious disease of rice worldwide. About thirty bacterial blight resistance gene in rice have been identified, five of them, Xa1, xa5, Xa21, Xa26 and Xa27 have been cloned successfully by a map based cloning approach. Xal4 is a dominant resistance gene against Philippines race 5 of bacterial blight pathogen Xoo, Taura et al. (1987) located it to the distal end on the long arm of chromosome 4, Tang et al (1998) located it between marker RG620 and G282 by RFLP markers, Bao et al. (2004) constructed a high-resolution linkage maps for Xal4 region by SSR markers, and located it between marker HZR970-8 and HZR988-1. In this paper, we had fine mapping Xal4 farther and analysis the sequence structure, expression and function of the resistance gene analogs in the Xa14 region.1. F₂ population consisting of 3300 individuals derived from a cross between IR24 and IRBB14 were screened for recombination events by 14 pair polymorphous SSR markers. Xal4 was located in a 324 kb region, flanked by the markers HZR970-8 and HZR669-2, cosegregated with HZR645-4, covered with nipponbare BAC clones OSJNBb0085C12 and OSJNBa0053K19 completely, OSJNBa0058K23 and BAC OSJNBB0060E08 partly.2. FGENESH and GENES CAN was used to analysis the nipponbare chromosome 4 DNA sequence of markers HZR970-8 and HZR988-1, a BLASTX analysis tool, CCD v2.06 was used to predict protein structure and function. BLASTX analysis showed that only 6 predicted resistance gene analogs had sequence homology with known plant R gene. They all encoded NBS-LRR proteins and clustered together to spread on nipponbare BAC OSJNBa0058K12, This 6 predicted genes were designated RGA1, RGA5, GRA6, RGA7, RGA12 and RGA14.3. Extract total RNA from the undamaged fresh leaves of IR24 and IRBB14 inoculated with PXO112 or water to investingate the expression of the six resistance gene analogs by RT-PCR, result showed RGA1 and RGA5 expressed in two parents, RGA12 only expressed in the resistant parent IRBB14. Extract total RNA from the undamaged fresh leaves of IR24, IRBB14, TP309 and MDJ, RT-PCR analysis indicated that RGA1 and RGA5 had constitutively expressed in four rice materials, RGA12 hadn't expressed in IR24, but constitutively expressed in other three rice materials. So we selected RGA1, RGA5 and RGA12 as the candidate genes of Xa14. The 3' ends of the...