一种适用于批量筛选和鉴定拟南芥突变体的快速DNA提取方法  

A rapid DNA extraction method for batch screening and identification of Arabidopsis mutants

作　　者：张小明[1,2] 勾雪娇[1,2] 梁鹏 林华明 赵晋洁 胡向阳[1] 

机构地区：[1]中国科学院昆明植物研究所西南野生生物种质资源库,云南昆明650201 [2]中国科学院大学,北京100049 [3]深圳华大基因科技服务有限公司,广东深圳518083 [4]广州基迪奥生物科技有限公司,广东广州510224

出　　处：《云南大学学报（自然科学版）》2015年第6期909-915,共7页Journal of Yunnan University(Natural Sciences Edition)

基　　金：国家自然科学基金(31470348)

摘　　要：DNA提取是拟南芥突变体筛选与鉴定中的关键环节.结合拟南芥突变体鉴定特点和DNA提取特点,对传统SDS法进行改良,摸索出一种简易、快速且经济的DNA提取方法,并将其应用到突变体的批量筛选与鉴定中.使用该方法可在常温下利用10 mg左右拟南芥叶片,以少量试剂,通过简单的研磨、离心、沉降和干燥等步骤便能快速制备用于PCR检测的DNA模板.采用本方法个人提取单个样品DNA仅需15 min左右,每小时可提取48个DNA样品.研究采用该方法提取了一批待鉴定拟南芥突变体的DNA样品,利用三引物法对其进行PCR检测,结果表明DNA样品浓度高,DNA扩增成功率高达100%,且目的条带清晰,说明该方法在拟南芥突变体的筛选和鉴定上是适用且高效的,可广泛应用于实验室突变体库的建立.DNA extraction is key to the screening and identification process of Arabidopsis mutants. Based on the characteristics of Arabidopsis mutants identification and DNA extraction,a simple,fast and economical DNA extraction method was worked out and applied into the batch screening and identification of mutants through modifying the traditional SDS extraction method. By using this method,DNA templates for PCR testing could be prepared quickly at room temperature from about 10 mg leaves,a small amount of reagent,and easy operation steps of grinding,centrifugation,sedimentation,and drying. A single DNA sample can be obtained in only about 15 minutes and 48 DNA samples are able to be extracted in an hour by the method. In this study,DNA samples of a batch of Arabidopsis mutants to be identified were prepared by applying the method,and then was detected by three primer PCR methods. The results showed that the concerntration of DNA samples was high,the success rate of DNA amplification was up to 100% and the target bands were clear and ideal,which indicates that the method is suitable and efficient,and can be widely used for mutant library constructions in the laboratory.

关 键 词：拟南芥 DNA提取 SDS抽提法 聚合酶链式反应 突变体鉴定 三引物法 

分 类 号：Q94-336[生物学—植物学]