琼脂糖电泳凝胶中回收微量DNA片段的新方法  

New and Simple Method for Recovery of Trace DNA Fragments from Normal Agarose Gel

作　　者：谭艳平[1] 杨玖英[1] 朱英国[2] 刘学群[1] 王春台[1] 

机构地区：[1]中南民族大学化学与生命科学学院生物技术国家民委重点实验室,武汉430074 [2]武汉大学生命科学学院植物发育生物学教育部重点实验室,武汉430072

出　　处：《中南民族大学学报（自然科学版）》2004年第1期10-14,共5页Journal of South-Central University for Nationalities：Natural Science Edition

基　　金：国家 973计划项目 (2 0 0 1 CB1 0 880 6 );湖北省重点科技研究资助项目 (992 T0 4 0 7)

摘　　要：对现有的 DNA片段回收方法进行了改进 .琼脂糖凝胶中目的 DNA片段条带切割后 ,浸入适量的 TE溶液 (0 .2～ 0 .4 m L/ g琼脂糖胶 ) ,在 37℃水浴保温 1 5 min,以溶出凝胶中的 DNA,利用相应的引物对浸出的微量DNA样品进行与原 PCR反应相同条件下的 PCR扩增 ,直接用 75 %乙醇沉淀后 ,适量 TE溶解得到回收的目的DNA样品 .此法回收的 DNA片段可用于分子克隆 ,特别是适用于 TA载体的克隆与测序 ,适合大批量微量样品的准确回收 ,防止了因试剂盒回收造成的微量To recover trace DNA fragments from the normal agarose gel after electrophoresis, a simple and economical retrieving procedure with high efficiency was designed. After the target DNA bands were cut from agarose gel, the DNA in the agarose slice was dissolved with appropriate dose of TE solution (0.2～0.4 mL/g gel) at 37 ℃ water bath for 15 min. The mixture was centrifuged at 12 000 g for 1 min, and 2~5μL supernatant was used for PCR amplification of the target DNA fragments with original primers under same conditions. Finally, the products were precipitated with 75 % ethanol and fragments with this method were suitable for use in molecular cloning, especially for cloning use with TA cloning kits.The new procedure makes it possible to recover a number of different fragments simultaneously, and protects the trace DNA losing in the procedure of DNA recovery as well.

关 键 词：琼脂糖凝胶电泳 DNA片段回收 正交实验法 聚合酶链式反应 

分 类 号：Q341[生物学—遗传学]