Cloning anS-RNaseallele, including the longest intron, from cultivars of European pear (Pyrus communisL.)

SummaryMost cultivars of European pear (Pyrus communis L.) exhibit S-RNase-based self-incompatibility. Their S-genotypes have mainly been assigned by analysing PCR-amplified S-RNase alleles from the genomic DNA of each cultivar. In this study, we identified eight European pear cultivars that had only one S-RNase allele amplified by conventional genomic PCR using a consensus primer pair. Rapid amplification of cDNA ends (RACE) on stylar RNAs of the cultivars was used to obtain the full-length sequence of the putative S 25-RNase. Long-PCR successfully amplified the S25-RNase allele, including a long 3,131 bp intron, the longest intron reported so far among S-RNase alleles in fruit tree species in the sub-tribe Pyrinae. Conventional PCR using a consensus primer and a primer designed from the intron sequence amplified a 385 bp fragment of the S25-RNase allele from the genomic DNA of all eight cultivars. An S-RNase-based, cleaved amplified polymorphic sequence (CAPS) marker system and S25-RNase allele-specific...