Abstract: In this study, ISSR molecular marker techniques were used to study genetic diversity and the genetic relationship of 35 Hovenia acerba germplasms from 12 provenances in Fujian. The reaction system of ISSR for analyzing the genetic diversity of H. acerba was established and optimized. The best one was shown as follows: total reaction volume 25 μL, 10×Ex Taq Buffer (Mg2+ plus) 2.5 μL, dNTP Mixture 375 μmol·L-1, primer 0.4 μmol·L-1, DNA template 75 ng, Ex Taq enzyme 1 U, ddH2O 14.55 μL. Procedures: pre-denaturation at 94 ℃ for 5 min, denaturation at 94 ℃ for 30 s, annealing at 50 ℃ for 1 min, extension to 72 ℃ for 90 s, a total of 35 cycles, final extension to 72 ℃ for 7 min. Finally, 175 locus of genomic DNA in total were detected from 10 ISSR primers and 155 were polymorphic locus, which covered 88.57%. The number of alleles(Na) was 1.8857 and the effective number of alleles(Ne) was 1.4300. Also, Nei’s gene diversity(H) was 0.2572 and Shannon’s information index(I) was 0.3959. UPGMA cluster analysis showed that according to the similarity coefficient of 0.74, the 35 copies of H. acerba germplasms could be classified into 4 groups. ISSR molecular marker techniques could well reveal genetic differences and the genetic diversity among H. acerba. The measured materials from 12 provenances of H. acerba showed abundant genetic diversity, which had great potential for exploitation and utilization.

Key words: Fujian province, Hovenia acerba, ISSR, reaction system, genetic diversity

摘要： 通过建立和优化枳椇Hovenia acerba的ISSR反应体系，并利用ISSR分子标记方法，对福建省12个种源地35份枳椇材料进行遗传多样性及亲缘关系研究。枳椇遗传多样性分析的ISSR最佳反应体系为反应总体积25 μL，10×Ex Taq Buffer(Mg2+ plus) 2.5 μL，dNTP Mixture 375 μmol·L-1，引物0.4 μmol·L-1，DNA模板75 ng，Ex Taq酶1 U，ddH2O 14.55 μL。程序：预变性94 ℃ 5 min；变性 94 ℃ 30 s，退火50 ℃ 1 min，延伸72 ℃ 90 s，共 35 个循环；最后延伸72 ℃ 7 min。利用筛选出的10个ISSR引物共检测了基因组DNA中175个位点，其中多态性位点155个，占总位点数的88.57%。供试材料观测等位基因数(Na) 1.8857、有效等位基因数(Ne) 1.4300，Nei’s基因多样性指数(H) 0.2572，Shannon多样性指数(I) 0.3959。UPGMA聚类分析表明，以遗传相似系数0.74为阈值，可将35份枳椇分成四大组。ISSR标记有效地揭示福建省枳椇种质资源丰富的遗传多样性，具较大开发利用价值。

关键词: 福建, 枳椇, ISSR, 反应体系, 遗传多样性