天牛幼虫保存与DNA提取方法比较研究  

Comparative studies on DNA extraction and preservation of Monochamus alternatus larva specimens

作　　者：郑斯竹[1] 安榆林[2] 徐梅[2] 杨晓军[2] 常虹[1] 嵇保中[1] 

机构地区：[1]南京林业大学森林资源与环境学院,江苏南京210037 [2]江苏出入境检验检疫局植物检疫实验室,江苏南京210001

出　　处：《中南林业科技大学学报》2012年第5期144-148,共5页Journal of Central South University of Forestry & Technology

基　　金：科技部国际合作项目(2009DFA31950);国家科技部质检总局项目(2011IK164);国家"十二五"科技支撑计划课题(2012BAK11B03)

摘　　要：为了开展天牛科昆虫分子系统学研究,采用酚-氯仿提取法、TakaraDNA快速提取试剂盒及GenMagBio动物细胞组织/细胞基因组DNA磁珠提取试剂盒3种方法分别对75%乙醇常温保存、无水乙醇-20℃冷藏保存和活体4℃保存3种保存方式且保存时间1年以上的松墨天牛幼虫进行DNA提取,并对不同提取方法所获取的DNA纯度与质量进行分析比较,确定了长时间保存天牛幼虫以无水乙醇浸渍-20℃冷藏保存效果最佳,而用GenMagBio磁珠法提取试剂盒提取DNA出现降解的标本提取效果最好。应用GenMagBio磁珠提取试剂盒对实验室保存多年的天牛幼虫标本进行DNA提取,并对提取DNA是否适合后续的PCR扩增及DNA序列测定进行了实验验证,结果符合预期。To explore the molecular phylogenetic relationship of Cerambycidae, the genome DNA of Monochamus alternatus larvae which were frozen under -20℃ , preserved in 100% ethanol or kept in 75% ethanol under normal temperature or kept as a living under 4℃ for more than one year, were extracted by using three methods, henol-chloroform method, Takara kit and GenMagBio kit. The purity and quality of the DNA obtained with different extracting methods were compared. The experimental results show that the preservation of the larvae stored in alcohol at -20℃ was the best way for the DNA extraction when the larvae must be preserved for a long time, and the extraction method of using GenMagBio Kit was much better than the other two methods, especially the DNA of larvae were breaking down. It was also proved that the DNA obtained were suitable for the subsequent PCR amplification and DNA sequence testing.

关 键 词：松墨天牛 天牛幼虫 保存方式 DNA提取 磁珠法 

分 类 号：S763.3[农业科学—森林保护学] Q781[农业科学—林学]