Résumé
Objective In this paper, the genetic diversity of 64 samples of Tetrastigma hemsleyanum germplasm resources in Chinese was analyzed. Methods ISSR-PCR was firstly used to amplify, and then POPGENE 32 software and NTSYS software was used to analyze the genetic diversity and phylogenetic relationship of 64 samples of T. hemsleyanum germplasm resources, and phylogenetic tree was constructed according to the UPGMA method. Results Ten primers with clear and reproducible bands were screened from 30 primers and used for genomic DNA amplification of 64 sample materials. A total of 83 polymorphic locis were amplified, whose polymorphic percentages were 71.43%-100% and average polymorphism percentage was 94.31%. The amplification polymorphic locis of primer S17 were the most (11) and the amplification polymorphic locis of primer P6 were the least (5), the average amplified polymorphic locis of 10 primers were 8.3. Genetic diversity analysis showed that the average number of alleles (Na) of 64 samples was 1.943 1, the average effective allele number (Ne) was 1.381 08, the average Nei’s gene diversity index (H) was 0.242 98, and the average Shannon diversity index (I) was 0.385 83. The variation range of the genetic similarity coefficient of the 64 samples was 0.431 8-0.988 6. A total of 64 samples were divided to six groups by UPGMA clustering method according to the similarity coefficient matrix when the genetic similarity coefficient was 0.715 5, which showed the abundant genetic diversity and relative gene stability of T. hemsleyanum germplasm resources. In addition, amplification figures by primers ISSR20, UBC857, and S17 were screened based on amplification result of 10 ISSR primers, and DNA fingerprinting was constructed, which can be used to identify 64 samples of T. hemsleyanum tested. Conclusion There are abundant genetic diversity and relative gene stability in T. hemsleyanum germplasm esources in China. ISSR analysis can reveal the genetic relationship among T. hemsleyanum germplasm resources in China, and provide certain reference for the evaluation, identification and new variety breeding of T. hemsleyanum germplasm resources in China.
Résumé
Objective: To establish an optimum reaction system suitable for ISSR analysis of Astragalus membranaceus and to analyze the genetic diversity of wild populations in Inner Mongolia. Methods: A stable and reliable ISSR reaction system was set up combining the concentration gradient of the single factor test and orthogonal test. The genetic diversity of 30 A. membranaceus populations in nine zones of Inner Mongolia was analyzed using NTSYS2.1 software. Results: The optimal ISSR reaction system (20 μL) contained 10 × PCR buffer 2.0 μL, 1.5 mmol/L MgCl2, 0.4 mmol/L deoxyribonucleotide triphosphate (dNTP), 1.5 U Taq DNA polymerase, 0.5 μmol/L primer, and 40 ng template DNA. A total of 169 amplified loci were detected by 15 ISSR primers, in which 157 loci were polymorphic loci with the percentage of 75%~100%. The genetic distance amplitude ranged between 0.242 7-0.730 8. The clustering analysis showed that 30 A. membranaceus populations could be divided into two categories, and most of them corresponded to the geographical distribution. Conclusion: ISSR-PCR reaction system for A. membranaceus is stable and reliable. Wild resources of A. membranaceus in Inner Mongolia have higher genetic diversity. The genetic relationship of the populations is correlated with its geographic location.