Isolation of S-locus F-box alleles in Prunus avium and their application in a novel method to determine self-incompatibility genotype.

2006 
This study characterises a series of 12 S-locus haplotype-specific F-box protein genes (SFB) in cherry (Prunus avium) that are likely candidates for the pollen component of gametophytic self-incompatibility in this species. Primers were designed to amplify 12 SFB alleles, including the introns present in the 5′ untranslated region; sequences representing the S-alleles S 1 , S 2 , S 3 , S 4 , S 4 ′, S 5 , S 6 , S 7 , S 10 , S 12 , S 13 and S 16 were cloned and characterized. [The nucleotide sequences reported in this paper have been submitted to the EMBL/GenBank database under the following accession numbers: PaSFB 1 (AY805048), PaSFB 2 (AY805049), PaSFB 3 (AY805057), PaSFB 4 (AY649872), PaSFB 4 ′ (AY649873), PaSFB 5 (AY805050), PaSFB 6 (AY805051), PaSFB 7 (AY805052), PaSFB 10 (AY805053), PaSFB 12 (AY805054), PaSFB 13 (AY805055), PaSFB 16 (AY805056).] Though the coding regions of six of these alleles have been reported previously, the intron sequence has previously been reported only for S 6 . Analysis of the introns revealed sequence and length polymorphisms. A novel, PCR-based method to genotype cultivars and wild accessions was developed which combines fluorescently labelled primers amplifying the intron of SFB with similar primers for the first intron of S-RNase alleles. Intron length polymorphisms were then ascertained using a semi-automated sequencer. The convenience and reliability of this method for the determination of the self-incompatibility (SI) genotype was demonstrated both in sweet cherry cultivars representing alleles S 1 to S 16 and in individuals from a wild population encompassing S-alleles S 17 to S 22 . This method will greatly expedite SI characterisation in sweet cherry and also facilitate large-scale studies of self-incompatibility in wild cherry and other Prunus populations.
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