摘要
以石杉科新鲜植物为材料,应用改良的CTAB法提取其基因组DNA,通过单因子实验优化了RAPD的反应体系。在25μL反应体系中,Mg2+2.0 mmol/L,dNTPs 0.3 mmol/L,模板DNA3ng/μL,随机引物0.2μmol/L,Taq DNA聚合酶1.25U。PCR循环程序为:94℃预变性5 min;然后94℃变性1 min,39℃退火1 min,72℃延伸1.5 min,36个循环;最后72℃延伸7 min,4℃保存。结果表明该体系具有良好的稳定性和重复性,可应用于石杉科植物系统分类,种的鉴定和遗传多样性研究。
DNA of Huperziaceae plant was extracted from the fresh leaves by using improved CTAB. The optimized reaction s ystem and programme of RAPD for Huperziaceae plant are as follows: The reactions were performed in a volume of 25 uL containing 2.0 mmol/L Mg^2+,0.3mmol/L dNTPs,3 ng/uL DNA template,0.2 umol/L random primer S8, 1.25 U Taq DNA polymerase. Thermal cycling was programmed by 1 cycle for S minutes at 94℃, then followed by 36 cycles for 1 minute at 94℃, 1 minute at 39℃, 1.5 minutes at 72℃, and finally by 1 cycle for 7 minutes at 72℃ ,storing at 4℃. The results show that this system is stable and repeatable and can be used in the studies of systematic classification of Huperziaceae plant, identification of variety and genetic diversity.
出处
《三明学院学报》
2009年第2期199-202,共4页
Journal of Sanming University
基金
福建省自然科学基金计划资助项目(2008J0301)
福建省科技厅重点项目(2006F6002)