摘要
目的对影响金钗石斛随机扩增多态DNA（RAPD）-PCR扩增效果的一些因素进行优化，为进一步研究其遗传多样性奠定基础。方法对金钗石斛RAPD分析中一些重要影响因素进行比较系统的构建和优化，建立金钗石斛RAPD稳定可靠的反应体系。结果金钗石斛较适宜的RAPD-PCR反应条件为：10μlPCR反应体系中，5-20ng模板DNA，3．0mmol·L^-1Mg^2＋，0．34mmol·L^-1dNTPs，0．6U Taq DNA聚合酶，1．5μmol·L^-1引物，较适宜的退火温度为36℃。结论所建立的金钗石斛RAPD反应体系具有标记位点清晰、反应系统稳定、检测多态性能力较强等特点，可用于金钗石斛的遗传多样性研究。以该优化的RAPD条件进行重复性实验，其实验结果重复性良好。
Objective To establish and optimize RAPD-PCR system of D. nobile Lindl. Methods The reliable RAPD - PCR system for the genetic diversity study of D. nobile Lindl. established by systematically testing and optimizing the important concentrations of the factors. Results The conditions that were suitable for RAPD - PCR of D. nobile Lindl. were as follows ： 10μl PCR reaction system contained 5 - 20 ng template DNA, 3.0 mmol · L ^-1Mg^2 ＋ , 0.34 mmol · L^- 1 dNTPs, 0.6 U Taq DNA polymerase and 1.5μmol · L^-lprime. The suitable annealing temperature in the RAPD -PCR reaction system was 36 ℃. Conclusion The RAPD -PCR system, which was established in this paper for studying D. nobile Lindl. , can provide clear, reliable, abundant polymorphic molecular markers, stable reaction system, and is suitable for studying the genetic diversity of D. nobile Lindl.. The experiment results are reproducible based on this conditions.
出处
《时珍国医国药》
CAS
CSCD
北大核心
2008年第12期2827-2830,共4页
Lishizhen Medicine and Materia Medica Research
基金
国家自然科学基金(No.30472150)
中国APEC科技产业合作基金项目
上海市重点学科建设项目资助(No.Y0301)
