Abstract
Nine full-length cDNAs of S ribonucleases (S-RNases) were cloned from stylar RNA of European pear cultivars by RT-PCR and 3′ and 5′ RACE. Comparison of the nucleotide sequences between the nine S-RNases cloned and 13 putative S alleles previously amplified by genomic PCRs revealed that seven corresponded to Sa, Sb, Sd, Se, Sh, Sk and Sl alleles, and the other two were new S alleles (designated as Sq and Sr alleles). Genomic PCR with a set of ȁ8FTQQYQȁ9 and ȁ8EP-anti-IIWPNVȁ9 primers was used to amplify nine S alleles; 1,414 bp (Sl), ca. 1.3 kb (Sk and Sq), 998 bp (Se), 440 bp (Sb) and ca. 350 bp (Sa, Sd, Sh and Sr). Among these, S alleles of similar size were discriminated by digestion with BaeI, BglII, BssHII, HindIII, EcoO109I and SphI. The PCR amplification of S allele following digestion with the restriction enzymes provided a PCR-RFLP system for rapid S-genotyping European pear cultivars harboring nine S alleles. The PCR-RFLP system assigned a total of 63 European pear cultivars to 25 genotypes. Among these, 14 genotypes were shared by two or more cultivars, which were cross-incompatible. These results suggested that the genes cloned represented the S-RNases from European pear, and that there were many cross-incompatible combinations among European pear varieties.
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Acknowledgements
This work was supported in part by Grants-in-Aid for Young Scientists (B) (no. 15780025), and Scientific Research (B) (no. 16380028) and (C) (no. 17580027) from the Ministry of Education, Science, Culture, Sports and Technology, Japan.
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Takasaki, T., Moriya, Y., Okada, K. et al. cDNA cloning of nine S alleles and establishment of a PCR-RFLP system for genotyping European pear cultivars. Theor Appl Genet 112, 1543–1552 (2006). https://doi.org/10.1007/s00122-006-0257-7
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DOI: https://doi.org/10.1007/s00122-006-0257-7