Study On Mechanism Of Self-Incompatibility In Chinese Plum (Prunus Salicina) And Self-Compatibility In Chinese Cherry (P.Pseudocerasus)

Using Chinese traditional plum cultivar, Chinese cherry cultivar 'Nanjing Chuisi' and its selfed progenies, we firstly identified their self-(in)compatibility, and then identified their S determinants as well as their S genotypes to studied on their mechanism of self-(in)compatibility. The results are shared as below:1. Experiments had been done to select appropriate method for suspending Chinese plum pollen grains. Pollen grain numbers and pollen germination ratio of thirty cultivars or strains had been investigated. Average distance cluster and discrimination analysis had been made on pollen grain numbers and pollen germination ratio of thirty cultivars or strains using software SAS8.2. The limit value of every level of pollen grain numbers and pollen germination ratio had been calculated. It was turned out that 10% glucose solution was appropriate for suspending Chinese plum pollen grains, and that Chinese plum had fertile male gametes. According to pollen grain numbers, thirty cultivars or strains had been classified into four obvious clusters, i.e.,Ⅰbeing more than 32,361 per flower,Ⅱranging from 22,556 to 32,361 per flower,Ⅲranging from 9,559 to 22,555 per flower,Ⅳbeing less than 9,559 per flower. According to pollen germination ratio, thirty cultivars or strains had been classified into four obvious clusters, i.e.,Ⅰbeing more than 75%,Ⅱranging from 75% to 52%,Ⅲranging from 51% to 25%,Ⅳbeing less than 25%.2. Natural self pollination have been done on 32 Chinese plum cultivars which fruit sets were caculated just before havest, artificial self pollination had been done on 5 cultivars, artificial cross pollination had been conducted on 16 cross combinations, pollen tube growthes in the style at 72 h after artificial self/cross pollination had been observed, fruit set were caculated at different time after artificial self/cross pollination. It was turned out that the fruit sets of all cultivars at havest after natural/artifcial self pollination were O demonstrating Chinese plum exhibiting self-incompatibility, and that fruit set of "Gaixiandali×Furongli" was O at havest after artificial cross pollination showing Chinese plum having cross-incompatible combination, and that "Gaixiandali×Huangpili" and "Jinshali×Younaili" which fruit sets were 3.15% and 4.67% respectifely at havest after artificial cross pollination showing Chinese plum having week-cross-incompatible combinations, and that there were 13 cross combinations which fruit sets were 7.76-22.90% at havest after artificial cross pollination showing Chinese plum having cross-compatible combinations, and that at 72 h after pollination, the pollen tubes could growth at the base of the styles in cross-compatible combinations, but could not in selfing and in cross-incompatible combination. All of above results demonstrated that Chinese plum exhibited classic gametophytic self-incompatibility. Furthermore, fruits had fallen till 40 d after pollination because of self-incompatibility in Chinese plum.3. Eight new genes, which mach necessary conditions for pistil S-determinant and were named as S-RNase-7～S-RNase-14 (Accession No. is AY71290, AY902455, AY996051, DQ003310, DQ512908, DQ512909, DQ512910 and EF177345 respectively), were identified in 10 Chinese traditional plum cultivars by PCR using S-RNase gene specific consensus primers. Furthrumore, S-RNase genotypes of 25 Chinese plum cultivars in all were determined. Abudent S-RNases diversity was identified in Chinese traditional plum cultivars. Fruit sets of fully compatible cross-combination were not always higher than that of semi-compatible cross-combinations. S-RNase-h had the highest gene frequency in sample of the present study implying its lingkage with some excellent agricultural traits.4. We determined eight novel sequences, which showed sequence high similarity to SFB genes of other Prunus species from 76% (between SFB-c from P. salicina and SFB-2 from P. armeniaca) to 87% (between SFB-7 from P. salicina and SFB-1 from P. mume), and named these sequences PsSFBs (P. salicina S-haplotype-specific F-box genes) (Accession No. is DQ849084, DQ849085, DQ849086, DQ849087, DQ849088, DQ849089 and DQ849090 respectively). The gene structure and the characteristic domains in deduced amino acid sequences of the PsSFB genes were similar to those of SFB genes from other Prunus. The eight identified PsSFB sequences showed S-haplotype-specific polymorphism with 74 to 83% amino acid sequence identity among them, and were exclusively expressed in the pollen. Three sequences from 410 bp to 2,800 bp of the intergenic region between the PsSFB sequences and the S-RNase alleles were obtained indicating their physical linkage. These results suggest that the PsSFB alleles are the pollen S-determinants of GSI in Chinese plum.5. Four PpsS-haplotypes (Prunus pseudocerasus S-haplotypes) comprising of at least two genes i.e., PpsS-RNase (P. pseudocerasus S-RNase) and PpsSFB (P. pseudocerasus S-haplotype-specific F-box), have been isolated in self-compatible tetraploid P. pseudocerasus CV. Nanjing Chuisi and its selfed progenies. S-genotype of 'Nanjing Chuisi' was determined as S-1/S-3'/S-5/S-7. These PpsS-RNases and PpsSFBs, respectively, expressed exclusively in style and in pollen, share the typical structure features with S-RNases and SFBs from other Prunus species, match with the necessary conditions for S-determinants, and comply with tetrosomic inheritence. SFB in PpsS-haplotype-3'was not amplified by PCR, while no mutations leading to dysfunction were found in S-RNase-3 and other S-haplotype based on their full-length c-DNAs. After self pollinantion, diploid pollen grains with S-genotypes S-7/S-1, S-7/S-5 and S-1/S-5 exhibited self-compatibility, while pollen grains with S-genotypes S-3'/S-7, S-3'/S-1 and S-3'/S-5 exhibited self-incompatibility. These results suggest that the PpsS-haplotype-1, PpsS-haplotype-5 and PpsS-haplotype-7 are functional, and that competitive interaction between two of them confer self-compatibility on cultivar 'Nanjing Chuisi'.