Comparison Of Genetic Diversity Of Pinellia Ternata (Thunb.) Berit Based On ISSR And SRAP

ObjectiveTo assess population genetic diversity of Pinellia ternata in different phenotype and different habitat by inter-simple sequence repeat (ISSR) and Sequence-related amplified polymorphism(SRAP).Methods1 To extract genomic DNA from Pinellia ternata.2 To compound thirty-six Pinellia ternata in different phenotype and different habitat and form different genomic pool.3 ISSR analysis:8 appropriate ISSR primers were selected from a total of 80 ISSR ones for ISSR PCR amplification.4 SRAP analysis:①Optimization for SRAP-PCR system of Pinellia ternata with orthogonal design and comprehensive scoring method.②14 appropriate SRAP primer pairs were selected from a total of 100 SRAP pairs for SRAP PCR amplification.Results1 ISSR analysis:①Cluster analysis of Pinellia ternata in different phenotype:A total 106 DNA bands and 87 polymorphic bands were amplified by 8 ISSR primers in Pinellia ternata and Pinellia pedatisecta Schott. The percentage of polymorphic bands was 82.1%. The number of polymorphic bands by an individual primer pair from 11 to 17, with a mean of 13.3. A total 88 DNA bands and 55 polymorphic bands were amplified in Pinellia ternata. The percentage of polymorphic bands was 62.5%. The number of polymorphic bands by an individual primer pair from 9 to 14, with a mean of 6.9. Jaccard's genetic similarity coefficients among Pinellia ternata in different phenotype based on the ISSR data ranged from 0.447 to 0.833.②Cluster analysis of Pinellia ternata in different habitat: A total 83 DNA bands and 65 polymorphic bands were amplified in Pinellia ternata. The percentage of polymorphic bands was 78.3%. The number of polymorphic bands by an individual primer pair from 7 to 16, with a mean of 10.4.③Cluster analysis of Pinellia ternata in different phenotype and different habitat:A total 104 DNA bands and 97 polymorphic bands were amplified in Pinellia ternata. The percentage of polymorphic bands was 93.3%. The number of polymorphic bands by an individual primer pair from 8 to 20, with a mean of 13.0.2 SRAP analysis:①A suitable SRAP reaction system was developed, i.e.1 x Taq polymerase reaction buffer,2.5mmol·L-1 Mg2+,1.0U Taq DNA polymerase,0.1 mmol·L-1 dNTPs,0.8μmol·L-1 primer and 50ng genomic DNA templates with total 25μL reaction solution.②Cluster analysis of Pinellia ternata in different phenotype:A total 259 DNA bands and 168 polymorphic bands were amplified by 14 SRAP primer pairs in Pinellia ternata and Pinellia pedatisecta Schott. The percentage of polymorphic bands was 64.9%. The number of polymorphic bands by an individual primer pair from 10 to 29, with a mean of 18.5. A total 224 DNA bands and 53 polymorphic bands were amplified in Pinellia ternata. The percentage of polymorphic bands was 23.7%. The number of polymorphic bands by an individual primer pair from 10 to 28, with a mean of 3.8. Jaccard's genetic similarity coefficients among Pinellia ternata in different phenotype based on the ISSR data ranged from 0.779 to 0.994.③Cluster analysis of Pinellia ternata in different habitat:A total 237 DNA bands and 33 polymorphic bands were amplified in Pinellia ternata. The percentage of polymorphic bands was 13.9%.The number of polymorphic bands by an individual primer pair from 12 to 29, with a mean of 16.9.④Cluster analysis of Pinellia ternata in different phenotype and different habitat:A total 171 DNA bands and 27 polymorphic bands were amplified in Pinellia ternata. The percentage of polymorphic bands was 15.8%. The number of polymorphic bands by an individual primer pair from 9 to 19, with a mean of 13.2.Conclusion1 ISSR analysis:①Results suggested that ISSR markers could be used as an effective molecular technique for the diversity study of Pinellia ternata.②The habitat was more important than the phenotype in identification of Pinellia ternata germplasm resource.2 SRAP analysis:①Results suggested that SRAP markers could be used as an effective molecular technique for the diversity study of Pinellia ternata.②The habitat was more important than the phenotype in identification of Pinellia ternata germplasm resource.3 The similar molecular cluster results were obtained from SRAP and ISSR analysis and the two types of markers showed different effective diversity detection scopes.