Resistance gene analogues of HcrVf2 in apple

The HcrVf2 gene, isolated from the Vf locus, is the first apple scab resistance gene cloned from apple. During the cloning of HcrVf2 it was found that the R-gene belongs to a cluster of four genes and that others HcrVf2-like genes are present in (unknown) loci of the apple genome. The genomic library of the apple cv. 'Florina' (Vf) used to identify HcrVf2 has been screened by hybridization using the whole HcrVf2 sequence as probe to identify all the BAC clones carrying sequences homologous to this gene. Eleven BAC clones not belonging to the contigs assembled to clone HcrVf2 were found. The relative overlapping of the 11 BACs was studied, an SSRs marker was developed from each contig and the SSRs were mapped in the cross 'Fiesta' × 'Discovery'. The analysis of six BAC clones was sufficient to identify all the HcrVf2-like sequences present on these 11 BAC clones. In addition to these BACs, other 7 clones, belonging to the contigs assembled to clone HcrVf2 and known to carrying (unstudied) HcrVf-like sequences, were also investigated. All the 13 BAC clones have been subcloned, subclones carrying HcrVf-like sequences identified, sequenced and for each HcrVf-like sequence a consensus sequence has been generated. Of these 13 BAC clones, 22 sequences showing homologies to HcrVf2 were retrieved. Nine sequences (including HcrVf4) encoded ORFs showing all seven domains found in HcrVf2, and six of them have been shown to be expressed constitutively in young scab uninfected apple leaves of 'Florina' by RT-PCR.