摘要
对党参基因组DNA提取方法优化、ISSR体系形成及引物筛选三方面进行探讨,为研究党参居群遗传多样性及药材DNA鉴定奠定基础。经比较改良CTAB法、改进的高盐SDS法和试剂盒三种常用DNA提取方法,发现改良CTAB法效果最佳;利用优化设计并结合有关文献优化ISSR反应体系,最优反应体系为:50μL总反应体积中含约20ng DNA模板,1.25UTaq DNA聚合酶,2.25mmol.L-1Mg2+,200μmol.L-1dNTP,0.50μmol.L-1引物。以此体系为基础进行引物筛选,在100条ISSR引物中筛选出13条扩增条带清晰、多态性较高、重复性好的引物。
To assess genetic diversity and to authenticate the medicinal materials of Codonopsis pilosula （ Franch. ） Nannf. the present work including DNA isolation, optimization of PCR assay of inter-simple sequence repeat （ISSR） and primers screening were investigated. Among three DNA isolation methods, improved CTAB, improved SDS and isolation kit, the improved CTAB was found to be the best. Based on priority selection design and those results in reported references, the optimal ISSR-PCR action was carried out in a volume of 50μL containing 20 ng of genomic DNA, 1.25 U of Taq polymerase, 1 × buffer, 200 μmol · L^-1 each of dATP, dGTP, dCTP and dTTP, 0.5 μmol · L^-1of primer, and 2.25 μmol · L^-1 Mg^2＋. According to this PCR system, thirteen of one hundred primers were chosen for their clarity, high polymorphism and repeatition.
出处
《植物研究》
CAS
CSCD
北大核心
2009年第3期346-351,共6页
Bulletin of Botanical Research
基金
西北农林科技大学引进人才基金项目
