樟树干叶DNA提取方法的研究

A Study on Methods for Extraction of DNA from Cinnamomum camphora Dry Leaves

利用硅胶干燥的樟树幼叶为材料，用改良的CTAB法进行DNA提取实验，并以樟树新鲜嫩叶为对比。结果表明，加入质量分数为3％可溶性PVP粉与材料充分研磨，在提取缓冲液中加入体积分数为2％的β-巯基乙醇，能有效地去除材料中的多酚类物质；用氯仿／异戊醇（24：1）提取裂解液2次所获得的上清，加入1／20体积5mol／L的NaCl，并用无水乙醇沉淀DNA，可以有效地去除多糖的干扰，所提取的DNA产率和纯度均较高。尽管干叶提取的DNA的平均产率（373ug／g）和纯度（A260／A280比值为1．54—1．92）比鲜叶提取的DNA的平均产率（467ug／g）和纯度（A260／A280比值为1．6—1．9）略低，但两者的RAPD—PCR扩增的条带都清晰、稳定、明亮，完整性好。

Cinnamomum camphora＇s genomic DNA from young buds and dried buds with silica gel was extracted by the modified CTAB method. The results showed that polyphenolics were effectively removed from materials by blendering 3% PVP to the ground materials under liquid nitrogen in mortar and acceding concentration of 2% β -mercaptoethanol in extracting buffer; the polysaccharides were effectively removed by adding 1/20 volume 5 mol/L sodium chloride solution and absolute ethanol to the blundering aqueous phase after suc- cessively extracting lysis solution twice with 24：1 chloroform： isoamyl alcohol; the high and purer DNA samples were obtained. Though the average yield （373 ug/g） and purity （A260/A280 between 1.54 - 1.92） of DNA extracted from dried buds were fewer than the DNA extracted from young buds, but the bands of RAPD - PCR amplified DNA were all clear, stable, bright and complete.