ISSR Analysis For Genetic Diversity Of Cypripedium In The Region Of Changbai Mountain

In this topic，ISSR molecular markers were used to analysis the genetic diversity ofCypripedium from Changbai Mountain. The objective was to comprehend the genetic diversityof Cypripedium. It was to provide scientific guidance and basis for protecting and utilizingCypripedium. The results were as follows.The optimal ISSR-PCR conditions for Cypripedium were established by single factorexperiment. The results show that containing of30ng template DNA，0.5μmol/L primer，0.2mmol/L dNTP，and0.75U Taq DNA polymerase in a volume of25μL was optimal.In this study，14ISSR primers were screened for the amplification of Cypripedium. Amongthe14ISSR primers，11ISSR primers were screened for the amplification of different colorCypripedium，94bands were resulted，of which86were polymorphic bands，and the percentageof polymorphic bands was91.5%.14ISSR primers were amplified for which the comparisonbetween Cypripedium and other three orchid，158bands were resulted，of which155werepolymorphic bands，the percentage of polymorphic bands was98.1%.14ISSR primers wereamplified for which the comparison between wild Cypripedium and introduction ofCypripedium，189bands were resulted，of which177were polymorphic bands，and thepercentage of polymorphic bands was93.7%.POPGENE analysis indicated that the genetic diversity of Cypripedium from ChangbaiMountain were relatively high. The genetic diversity of different color Cypripedium were that na=1.6364，ne=1.4182，He=0.2500，Ho=3697. Among the genetic diversity of the comparisonbetween Cypripedium and other3orchid，the genetic diversity of Cypripedium were that na=2.0000，ne=1.8993，He=0.4684，Ho=0.6606，the genetic diversity of Tulotis fuscescens andOreorchis patens were that na=2.0000，ne=1.5429，He=0.3519，Ho=0.5367，the geneticdiversity of Platanthera chlorantha were that na=2.0000，ne=1.9115，He=0.4768，Ho=0.6698. The genetic diversity of wild Cypripedium were that na=2.0000，ne=1.9461，He=0.4842，Ho=0.6769. The genetic diversity of introduction of Cypripedium were na=2.0000，ne=1.9567，He=0.4878，Ho=0.6806. No obvious difference in genetic diversity between wildCypripedium and introduction of Cypripedium.Genetic identity and genetic distance were accounted by POPGENE. It indicated that the genetic identity each other in11individuals of different color Cypripedium was in the range of0.4468to0.78720and the genetic distance was the range of0.2806to0.8056. The geneticidentity each other in Cypripedium and other3orchid was in the range of0.4684to0.9114andthe genetic distance was the range of0.0928to0.7585. The genetic identity and genetic distancebetween wild Cypripedium and introduction of Cypripedium was0.9444，0.0572.The result of UPGMA clustering analysis by NTSYS of different color Cypripediumshowed that the11individuals were divided into3categories. The first kind was the4colorsChypripedium acranthum Sw，the second kind was the2colors Cypripedium calceolus L andCypripedium X ventricosum Sw，the third kind was Cypripedium shanxiense.C.Chen and themodification of Cypripedium calceolus L. The UPGMA clustering analysis and primary axisanalysis were used to the wild Cypripedium and introduction of Cypripedium. It indicated that20individuals were divided into3categories. The first kind was Chypripedium acranthum Swand Cypripedium X ventricosum Sw，the second kind was Cypripedium calceolus L andCypripedium shanxiense.C.Chen，the third kind was Cypripedium guttatum.According to the genetic diversity of Cypripedium，the main reson for the rapid decrease ofCypripedium was caused by mankind. Mainly reflected in over-excavation and the destructionof the living environment. The awareness of protect the plant must be strengthed. The strategiesof protecting in its native environment and protecting in other environment could be took intoaccount.