白木香愈伤组织与叶片DNA快速提取方法的确定  

Determination of the Qualified Rapid-DNA-Extraction Method from Callus or Leaves of Aquilaria sinensis

作　　者：高志晖[1] 赵文婷[1,2] 张争[1,3] 徐艳红[1] 金钺[1] 杨云[3] 魏建和[1,3] 

机构地区：[1]中国医学科学院药用植物研究所,北京100193 [2]东北林业大学,黑龙江哈尔滨150040 [3]中国医学科学院药用植物研究所海南分所,海南省南药资源保护与开发重点实验室,海南万宁571533

出　　处：《安徽农业科学》2014年第14期4194-4196,共3页Journal of Anhui Agricultural Sciences

基　　金：国家自然科学基金(No.81001607;No.81173481);教育部新世纪优秀人才支持计划项目(2008)

摘　　要：[目的]确定白木香愈伤组织与叶片DNA的快速提取方法。[方法]用TE、ES1、ES2 3种提取液及相应的方法提取叶片和愈伤组织总DNA,通过琼脂糖凝胶电泳检测提取产物,通过普通PCR及荧光定量PCR检查DNA的可用性,并将PCR产物经克隆测序验证。[结果]ES1法提取的叶片和愈伤DNA条带清晰,纯度较高;普通PCR和荧光定量PCR可扩增出200和600 bp左右的目的条带。ES2法提取液电泳不能检测到DNA条带,但PCR可扩增出愈伤组织200 bp左右目的条带。TE法得到的提取液检测不到DNA条带且PCR不能得到目的条带。[结论]ES1提取液及相应的提取方法可用于白木香愈伤和叶片DNA高通量快速提取,为沉香属植物种质研究和分子研究奠定基础。[Objective] To determine the rapid DNA extraction method from calli or leaves of Aquilaria sinensis.[Method] We tested three methods （TE,ES1,ES2） used in Arabidopsis and rice for rapid DNA extraction from leaves and calli ofA.sinensis.The extracted products were evaluated through agarose gel electrophoresis,PCR and real-time PCR reactions.The amplicons were further confirmed by clone and sequencing.[Result] The extraction method using solution ES1 could successfully isolate total DNA from leaves and calli ofA.sinensis in 20 min：agarose gel electrophoresis showed clear binds.The DNA was qualified for PCR amplification of ～ 200 bp or ～ 600 bp sequences.The extracted product from calli using solution ES2 can also amplify the ～ 200 bp sequence,but failed to amplify the ～600 bp sequence.Other methods were failed to extract DNA and amplify any sequences.[Conclusion] The method using ES1 solution is qualified for rapid DNA isolation from A.sinensis for PCR amplification,which will lay a foundation for Aquilaria plants germplasm and molecular research.

关 键 词：濒危南药 白木香[Aquilaria SINENSIS (Lour ) Gilg] DNA快速提取 沉香 叶片 愈伤 

分 类 号：S567[农业科学—中草药栽培]