| This study identified the self-incompatibility S genotype of 40 pear varieties and studied their self-incompatibility characteristics by using pear as the experimental material and combining the techniques of genomic DNA specific PCR amplification,fragment recovery,cloning and sequencing.Through field pollination experiment and in situ germination detection of pollen tube,we found two self-compatible varieties Meigetsu and XinXue to study their affinity variation characteristics,which provided a possibility for the selection and breeding of new self-compatible varieties.The experimental results are as follows:1.Using the universal primers ’FTQQYQ’ and ’anti-IIWPNV’ of pear self-incompatibility genes,the S genotypes of 40 pear varieties were identified,including 6 P.pyrifolia varieties,7 P.bretschneideri varieties,5 P.communis varieties,7 interspecific hybrids,1 P.ussuriensis variety and 14 local varieties.A total of 22 different S.genotypes were identified:S1,S3,S5,S6,S12,S16,S17,S19,S20,S21,S22,S27,S35,S39,S59,S101,S116,S108,Sa,Sb,Se,etc.Among them,S19identified from ’Hongbeileisha’ was S gene of Pyrus betulifolia after comparative analysis.The full-length sequence of the S6-RNase gene was cloned from the Xinxue style RNA and registered in Genebank under the accession number(MG834536).After comparison analysis,Genebank has been registered in the S gene,S33-RNase(accession number is DQ138081.1)is the same gene as S6-RNase.2.In this study,the self-crossing fruit setting rate of Meigetsu pear was 39%,and the pollen tube could grow to the bottom of the style,which was considered as the self-compatibility variety.In order to study its mutation mechanism,we conducted self-cross-pollination and cross-pollination,and determined its mutation type by analyzing the separation rule of S genotype in the progeny.S3S3 and S4S4 genotypes appeared in the self-crossing progeny of Meigetsu,indicating that both S genotype were mutated.As female parent in hybrid combinations,Meigetsu ×Cuiguan progeny S3S3 and S3S4 two genotypes of 0,S3S5,S4S5 approximately 1:1 proportion of separation,Meigetsu×ZhongliNO.1 progeny S3S35 hybrid combinations,the individual number of S4S35 genotypes respectively,14,21 ratio is close to 1:1,accord with Mendel’s genetic law,thus speculate that Meigetsu style of S3 and S4 both S genotype to maintain normal function.The number of S3S3,S3S5,S3S4,and S4S5 genotypes in the progeny of Meigetsu,the hybrid combination of Cuiguan and Meigetsu as the male parent is 3,8,4,and 16,respectively,which is significantly different from the theoretical ratio of 1:1:1:1 indicating that the ’ Meigetsu’ pollen had mutation.The above studies prove that’Meigetsu’is caused by pollenmutation.However,the specific mutation mechanism of pollen has not been solved,and further studies in this regard are needed.3.The self-pollination setting rate of Xinxue reached 22.9%(more than 20%)and the pollen tube was observed at the bottom of the style,which was considered as the self-compatibility variety.In order to investigate the mutation mechanismofXinxue,we studied the genetic distribution of S5-and S6-RNase alleles in the self-pollinated progeny of Xinxue to detect the mutant Sgene.The results showed that S5S5 and S6S6 genotypes were detected in progeny,indicating that both S5-and S6-haplotypes were functionally abnormal.When Xinxue as female parent,S5S5and S5S6 genotypes were not found in the progeny of Xinxue×Cuiguan and S5S6 and S6S6were not found in the progeny of Xinxue×Jincunqiu.This indicated that the S5-and S6-RNase genes in the style of Xinxue were not mutated.However,the separation ratio of S3S5,S5S5,S3S6and S5S6 in the progeny of Cuiguan×Xinxue was close to 1:1:1:1,and the separation ratio of S6S6,S5S6,S5S22 and S6S22 in the progeny of Jiuqunachangba×Xinxue was also close to 1:1:1:1,which was in line with the theoretical ratio.It indicates that the Xinxue pollen mutates,but exactly how the pollen is mutated is unknown at present,which can only be determined after the S genotype of pear pollen is determined. |
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