海南龙血树ISSR-PCR反应体系建立与有效引物筛选

Establishment of ISSR-PCR Reaction System and Screening of Effective Primers with ISSR Marker for Dracaena cambodiana

摘要:

采用单因子试验与正交试验的方法, 研究了海南龙血树(Dracaena cambodiana Pierre ex Gagnep) ISSR-PCR反应体系中的主要影响因子, 筛选出16条有效引物并优化了它们的退火温度, 此外还检测了体系的重复性。优化后的25 μL反应体系包含：10× PCR buffer 2.5 μL, 3.0 mmol/L MgCl2, 300 μmol/L dNTPs, Taq聚合酶3.0 U, 引物0.2 μmol/L, DNA模板20 ng。该反应体系可用于海南龙血树的遗传多样性和遗传结构的分析。

Abstract:

The influence factors of ISSR-PCR for Dracaena cambodiana Pierre ex Gagnep were studied by using single factor test and orthogonal design, and a set of ISSR-PCR reaction system was established. Sixteen ISSR primers were selected out, and their annealing temperatures were also optimized. Furthermore, the optimized reaction conditions were examined for its repeatability in D. cambodiana. The optimum ISSR-PCR reaction system with total 25 μL reaction solution contain 2.5 μL 10×PCR buffer, 3.0 mmol/L MgCl2, 300 μmol/L dNTPs, 3.0 U Taq polymerase, 0.2 μmol/L primers, and 20 ng DNA template, which can be used for analysis in genetic diversity and genetic structure of D. cambodianas.