[Construction of a deep coverage rice BAC library and identification of clones associated with disease-resistant...

A BAC library for IRBB56, an accession pyramiding Xa4, xa5 and xa13 three bacterial blight resistance genes, was constructed. The library contains 55,296 clones with an average insert size of 132 kb. Based on a haploid genome size of 450 Mb, the coverage of the library was about 14 genome equivalents that make it one of the most comprehensive BAC libraries available in rice and provide 99.99% possibility to isolate any interested rice genes or sequences in the library. To determine the representation of organelle DNA homologues in the library, the library was screened with three different chloroplast genes and four mitochondrial genes, respectively. Results from this screening showed that less than 1% of clones in the library contain organelle genomic DNA homologues. Then, DNA markers on three different chromosomes linked to Xa4, xa5, and xa13, and a PCR fragment of rice UROD gene, were used to screen the library resulting in a range of 11-106 hits that will promote the isolation of these genes. The deep coverage and the large insert size of the library will facilitate physical mapping, isolation, and cloning of rice genes.