Genetic Diversity Of Two Geographical Populations Of Pelodiscus Sinensis Revealed By ISSR Analysis

Pelodiscus sinensis, is an economic reptile species in China.It has high edible and medicinal value. Understanding the levels of genetic diversity, genetic structure and the degree of genetic differentiation of Pelodiscus sinensis has an very important theoretical guidance for the evaluation and reasonable utilization of its natural resource. In this study, inter simple sequencer epeat (ISSR) molecular marker was applied to investigate the genetic diversity of two populations of Pelodiscus sinensis, including Huai River population and Taiwan population. The main results were as follows:1. Through the orthogonal experimental design, ISSR reaction conditions and reaction system for Pelodiscus sinensis were optimized. The optimal conditions of ISSR reaction were as follows:25μL reaction volume including0.4μmol/L ISSR primer,30ng DNA template,2.5mmol/L Mg2-,0.2mmol/L dNTPs,0.2U Taq DNA polymerase. The optimum ISSR reaction program for Pelodiscus sinensis was as follows:5min preamplification denaturation at94℃berore the first cycle,40cycles of40s at94℃,45s at a primer-specific annealing temperature(52-58℃), and60s at72℃,10min after the40cycles at72℃2.8ISSR primers were screened to amplify37individuals belonging to2geographical populations of Pelodiscus sinensis. A total of77reproducible DNA fragments were amplified,47fragments were polymorphic, and the percentage of polymorphic loci was61.04%. The percentages of polymorphic loci, Effective number of alleles, Nei’s gene diversity, Shannon’s information index were estimated by POPGENE software. The percentages of polymorphic loci of two populations were59.57%and51.06%; The effective number of alleles were1.5603and1.5222; The Nei’s gene diversity were0.1657and0.1524; The Shannon’s information index were0.2164and0.1977, respectively. The results indicated that the two populations had low genetic diversity, the Huai River population had higher genetic diversity comparatively.3. The Gst (Coefficient of genetic differentiation) according to the analysis of Nei’S gene diversity was0.0796. It showed medium genetic differentiation occurred between2populations. The Nm(Gene flow) based on Gst was5.7778, it showed In this research, the Nei’s unbiased genetic distances between2populations of Pelodiscus sinensis were0.0837(smaller than0.2), it showed that significant genetic difference among the2populations has formed. UPGMA cluster analysis based on Nei’s genetic distances showed that37individuals were divided into two clusters corresponding to two populations.