Standies On Mechanism Of Self-Compatibility And S-Alleles Evolution In Prunus Species

Chinese cherry (Prunus pseudocerasus), sweet cherry (P. avium), the flowering cherry (P. speciosa), apricot (P. armeniaca) and some of self- and cross-pollinated progenies were used as materials. Based on their self-compatibility, stylar and pollen S alleles and some of the S-genotypes were identified to study the mechanism of Chinese cherry and Prunus S-allele evolution. The results are as follows:1. Artificial self-pollination had been done on nine Chinese cherry cultivars. The pollen tubes growths in the styles were observed at 72 h after self-pollination, and the fruit sets were calculated before harvest. The results showed that nine Chiese cherry cultivars were self-compatible, which means that the pollen tubes could grow at the bottom of style at 72 h after self-pollination, suggesting that Chinese cherry were the fruit trees of gametophytic self-compatibility. We detected that these Chinese cherry were polyploid cultivars, and more than two S alleles are contained in cultivars.2. Ten different stylar S-RNase alleles were identified from fourteen cultivars by PCR and RT-PCR with conserved primers which were designed from Prunus S-RNases, S-genotypes were also identified in nine cultivars. The deduced amino acid identities ranged from 69.4 to 84.3% among ten S-RNases, and ranged from 65.31 to 100% with other Prunus S-RNases. Five S-RNases in Chinese cherry have exceptionally high identities with other Prunus S-RNases (more than 97%). The exceptionally high similarities were found between their first and second introns. Moreover, four SFB alleles were identified from seven Chinese cherry cultivars by RT-PCR with conserved primers designed from Prunus SFBs. They were specifically expressed in pollen and tightly linked with stylar S-RNases. The deduced amino acid identities ranged from 76.9 to 80.0% among them. The alignments showed that the exceptionally high identities were also found between Chinese cherry and other Prunus species SFBs. For example, the identities between PpsSFB1 and PmSLF9 and between PpsSFB5 and PspiSFB16 were more than 96%.3. The fruits could be harvested in cross-pollination between sweet cherry and Chinese cherry. Its S-genotypes were identified by PCR with conserved primers of S-RNase and SFB alleles, which could determine that the distributions and primary genetic model of S-haplotypes expressed in pollen grains of tetraploid Chinese cherry. The analysis of S-haplotypes in self- and cross-pollinated progenies of Chinese cherry cultivars showed that the self-compatibility of Chinese cherry was not resulted from accumulation of non-functional S-haplotypes, but was from competitive interaction. Moreover, the distributions of the S-haplotypes derived from the the same Chinese cherry cultivars were disequilibrium in self- and cross-pollinated progenies, which indicated that the compatibility between the different pollen grains and the same style and between the same pollen grains and the different style were not identical. These results suggested that the compatibility between the hetero-diploid pollen grains and the style or the growth of these pollen tubes were regulated by some factors transmitted from style or ovule.4. In order to investigate the evolutionary relationship between S alleles and Prunus species, eight S-RNases, eight SFBs and one SFB-like gene were cloned from two wild Chinese cherry accessions and the two flowering cherry. Four S-RNases and one SFB alleles had exceptionally high identities with other Prunus S-RNase and SFB alleles, respecitively. The results of the alignments and phylogenetic trees suggested supporting the hypothesis that S-RNase alleles were evolved before the divergence of Prunus species. When comparing the deduced amino acid identities of Prunus SFBs with S-RNases, it was obviously that the sequence polymorphism of SFBs was less than that of S-RNases, which indicated that the origin of SFB alleles was later than S-RNase alleles. And the distribution of SFBs was not consist with their S-RNases in phylogentic trees, indicated that the S-RNase and SFB alleles were separated but correlated during the co-evolutionary process.5. S4- and S24-haplotypes were cloned from Northern American apricot ’Hereot’ and Chinese apricot ’Chuanzhihong’. Their S-RNase and SFB alleles had exceptionally high identities, which were used to determine the recognition specificity of self-incompatibility. The fruit sets and the pollen tube growths showed that these two cultivars were self-incompatible and their cross-pollinations were compatible. The analysis of S-genotypes in their cross-pollinate progenies showed that the stylar S4-RNase and S24-RNase rejected the pollen grains of S4 and S24, which suggested that the two S-RNases and two SFBs in S4- and S24-haplotypes had the dual-specificity. Based on these conclusions, we proposed that dual-specificity was a crucial step in the evolutionary process without loss the self-incompatiblity.