Chromosome landing at the bacterial blight resistance gene Xa4 locus usinga deep coverage rice BAC library

Xa4 is a dominantly inherited rice gene that confers resistance to Philippi ne race I of the bacterial blight pathogen Xanthomonas oryzae pv. oryzae in rice. In order to isolate the gene by positional cloning, a bacterial arti ficial chromosome (BAC) library was constructed from genomic DNA isolated f rom an Xa4-harboring accession, IRBB56. The library contains 55,296 clones with an average insert size of 132 kb, providing 14 rice genome equivalents . Three DNA markers closely linked to Xa4 were used to screen the library. The marker RS13, a resistance gene analogue that co-segregates with Xa4, id entified 18 clones, of which four and six, respectively, were simultaneousl y detected by the other two markers. G181 and L1044. Fingerprinting and Sou thern analysis indicated that these clones overlapped and define an interva l spanning 420 kb. In an F-2 population derived from an indica variety, IR2 4, and its Xa4-containing near isogenic line (NIL). IRBB4, the susceptible plants were screened in order to map the Xa4 gene genetically and physicall y. Out of 24 insert ends isolated from the BACs in the contig, three reveal ed polymorphisms between IR24 and IRBB4. Two insert ends, 56M22F and 26D24R , flanked Xa4 on each side. Based on the overlap of the BACs, six overlappi ng clones were considered to include the Xa4 allele, one of which. 106P13, was chosen for further investigation.