Study Of Genetic Diversity Based On Molecular Marker In Germplasms Resources Of Dendrobium Officinale

D. officinale is rare Medicinal materials in China. To investigate the genetic diversity inpopulations of D. officinale, and provide the basis for the protection of germplasm resources andbreeding of new varieties. In this thesis, we used the SCoT and SSR markers to build SCoT andSSR-PCR reaction system, and analysed the genetic diversity of D. officinale. The results were asfollows:1．The optimum SCoT-PCR reaction system （20μL）for D. officinale was established: including DNAtemplate30ng, Mg²⁺2.5mmol·L^-1, dNTPs0.15mmol·L^-1,TaqDNA polymerase1.5U and primer0.4μmol·L^-1. The PCR amplifications followed a given protocol:5min at94℃,35cycles of35s at94℃,35s at51℃and1.5min at72℃, and at72℃for10min finally.The optimum reaction system wasfurther verified in32D. officinale varieties, which showed good stability and repeatability.Genetic diversity and cluster analysis were conducted among individual and population of D.officinale by SCoT. The results showed that28primers screened from57SCoT primers were used inthe SCoT-PCR amplification; And385bands were generated among which366bands werepolymorphic. The percentage of polymorphic bands （PPB） was94.67%; The percentage of polymorphicbands （PPL） for17populations of D. officinale ranged from28.83%to64.42%was45.73%（mean）.Cluster results showed that D. officinale from different source stayed together, With Some Exceptions,which showed that there were rich in genetic diversity.2．The optimum SCoT-PCR reaction system （20μL）for D. officinale was established: containing40ngDNA template,1.5mmol·L^-1Mg²⁺,0.40mmol·L^-1dNTPs,0.5U Taq DNA polymerase and0.6μmol·L^-1primer. The PCR amplifications followed a given protocol:5min at94℃for5min,35cycles of at95℃for35s, at （41⁵8.1）℃for35s and at72℃for1.5min, and extending at72℃for10min finally.The optimum reaction system was further verified in95D. officinale varieties, which showed goodstability and repeatability.Genetic diversity and cluster analysis were conducted among individual and population of D.officinale by SSR. The results showed that10pair of primers screened from26pair of SSR primerswere used in the PCR amplification; And63bands were generated among which61bands werepolymorphic. The percentage of polymorphic bands （PPB） was97.00%; The percentage of polymorphic bands （PPL） for17populations of D. officinale ranged from11.11%to90.48%was53.22%（mean）.The cluster analysis results showed that Distinct genetic differences and extensive genetic diversitywere presented among cultivated D. officinale populations.3. The genetic diversity of individual and populations of D. officinale were studied with SCoT and SSRmarkers together. The results showed that the genetic diversity of D. officinale is among cultivated D.officinale populations. Genetic relationship of D. officinale has a obvious correlation with its geographyprovenance. Cluster analysis showed SCoT and SSR markers was reliable with genetic diversity andgenetic relationship.This would provide the reliable theory basis for the variety breeding, introduction,and extension of D. officinale.