摘要
为了从富含次生代谢物的樟科植物肉桂、锡兰肉桂、阴香中获得高质量DNA ,研究和改进了CTAB法、高盐低pH法和尿素法。改进方法包括 :1)在裂解液中加入 2 % β -巯基乙醇和 5 %PVP ,以防止氧化褐变的发生 ;2 )在酚 :氯仿抽提前加入 1 5mol L醋酸铵冰浴处理 ,能降低DNA的黏性。所得DNA的质量和产量经电泳、紫外吸收A2 60 A2 80 、PCR扩增和限制性内切酶酶切检测 ,结果表明改进法提取的DNA质量要比常规法的好。其中改进的CTAB法获得的DNA纯度最高 ,能用于PCR扩增和限制性内切酶酶切 ,是提取这 3种樟科植物总DNA的最佳方法。
CTAB method,low pH medium with high salt method (LPHS),and urea method were used and improved to extract high-quality DNA from Lauraceae plant Cinnamomum cassia Presl.,C.zeylanicum and C.burmannii,which are rich in secondary metabolites.The modified protocols involved two key steps:1) adding 2% β-mercaptoethnoal and 5% polyvinylpyrrolidone (PVP) in extraction solution to inhibit the oxidization of polyphenols;2) treating with 1.5?mol/L NH 4Ac and incubation at 0℃ prior to supernatant extraction with phenol:chloroform to lower the viscosity of DNA.The yields and qualities of DNA were analyzed by electrophoresis,ratios of A (260)/A (280),PCR amplification and restriction digestion.The results showed that the modified methods are better than the original ones in terms of the quality of total DNA.Among them,the modified CTAB method,which can produce high quality DNA suitable for PCR amplification and restriction digestion,is the best one for DNA extraction from three species of Lauraceae.
出处
《云南植物研究》
CSCD
北大核心
2004年第4期451-457,共7页
Acta Botanica Yunnanica
基金
厦门大学预研基金资助
关键词
肉桂
锡兰肉桂
阴香
DNA提取
Cinnamomum cassia
Cinnamomum zeylanicum
Cinnamomum burmannii
DNA extraction
