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长白山杜鹃花基因组DNA提取及RAPD体系的建立
Establishment of RAPD System and DNA Extraction from Rhododendron in Changbai Moutain
【摘要】 采用改良的CTAB法提取杜鹃花的基因组DNA,所得的DNA纯度高、质量好,可用于RAPD分析。筛选出的杜鹃花RAPD反应的最佳体系为25μL反应体系中包括模板DNA20~200 ng,引物10pmol,dNTPs100μmol·L-1,TaqDNA聚合酶0.5U,Mg2+2.5 mmol·L-1,10×buffer缓冲液2.5 mmol·L-1,其余部分用无菌超纯水补充。PCR扩增程序为94℃预变性5 min;94℃变性1 min,37℃退火1min,72℃延伸2min,40次循环;72℃最终延伸7 min。应用优化后的反应体系PCR扩增获得的RAPD指纹图谱带型清晰,重复性好,为长白山杜鹃花品种鉴定、分类的研究提供了一定基础。
【Abstract】 High purity and quality DNA extracted from Rhododendron was used for RAPD analysis.The best RAPD reaction system optimized was: total volume 25μL,containing,contained 2 template DNA 10~200ng,primer 10pmol,dNTPs 100 μmol·L-1,MgCl2 2.5 mmol·L-1,Taq DNA polymerase 0.5unit,10×buffer 2.5 μL,left complement with dd H2O.PCR amplification condition was: pre-denaturation at 94℃ for 5min;Denaturation at 94℃ for 1min,annealing at 37℃ for 1min,extension at 72℃ for 2min,cycle for 40 times.Final extension at 72℃ for 7min.RAPD fingerprinting obtained by the optimized system was clear and repetitive,which provided a certain foundation for Rhododendron variety identification and category in Changbai Mountains.
- 【文献出处】 湖北农业科学 ,Hubei Agricultural Sciences , 编辑部邮箱 ,2008年03期
- 【分类号】S685.21
- 【被引频次】10
- 【下载频次】283