Construction of a 1.2-Mb contig including the citrus tristeza virus resistance gene locus using a bacterial artificial chromosome library of Poncirustrifoliata (L.) Raf.

The citrus tristeza virus resistance gene (Ctv) is a single dominant gene i n Poncirus trifoliata, a sexually compatible relative of citrus. To clone t his gene, a bacterial artificial chromosome (BAC) library has been construc ted from an individual plant that was homozygous for Ctv. This library cont ains 45 696 clones with an average insert size of 80 kb, corresponding to 9 .6 genome equivalents. Screening of the BAC library with five chloroplast D NA probes indicated that 0.58% of the BAC clones contained chloroplast-deri ved inserts. The chromosome walk across the Ctv locus was initiated using t hree closely linked genetic markers: C19, AD8, and Z16. The walk has been c ompleted and a contig of ca. 1.2 Mb was constructed. Based on new data, the genetic map in the Ctv region was revised, with Ctv being located between AD8-Z16 and C19 at distances of 1.2 and 0.6 cM, respectively. Utilizing DNA fragments isolated from the contig as RFLP markers, the Ctv locus was furt her mapped to a region of ca. 300 kb. This contig contains several putative disease-resistance genes similar to the rice Xa21 gene, the tomato Cf-2 ge ne, and the Arabidopsis thaliana RPS2 gene. This library will therefore all ow cloning of Ctv and other putative disease-resistance genes.