摘要
尝试利用全式金生物技术有限公司的TransDirect Animal Tissue PCR Kit^■试剂盒对整头茧蜂标本DNA进行提取,通过缩短浸泡时间和热激时间、优化取虫工具且试验后试虫经纯水漂洗2次以上,无损提取了实验室液浸标本及超过20年的针插干标本的DNA.使用微量分光光度计测定了所得的DNA样品在260和280 nm波长下的光密度值,验证了DNA浓度和质量.用28S、COⅠ、16S和EF-1α的4对引物对提取的DNA样品进行PCR扩增,PCR产物经琼脂糖凝胶电泳检测可得到单一、明显的条带,且通过测序进一步证明了序列的准确性.试验结果证明无损提取DNA的方法可行.
TransDirect Animal Tissue PCR Kit^■ from TransGen Biotech Co., Ltd. was used to extract the DNA of whole Braconidae specimens. With the assistance of hand-made insect picker, ethanol-preserved and dry specimens were soaked in the buffer solution and underwent heat shock briefly. After rinsing twice in pure water, the specimens DNAs were non-destructively extracted. Subsequently, optical density at 260 and 280 nm was measured with a micro-spectrophotometer for DNA quantity and quality assessment. PCR electrophoresis of DNA samples amplified by 28S rDNA, COⅠ, 16S rDNA and EF-1α gene produced bright and single bands at expected sizes, and accuracies of the sequences were further verified by sequencing technology. Experimental results showed that this DNA extraction with non-destructive method is feasible.
作者
夏慧敏
黄珺
郑敏琳
陈湜
陈家骅
XIA Huimin;HUANG Jun;ZHENG Minlin;CHEN Shi;CHEN Jiahua(Institute of Beneficial Insects,College of Plant Protection,Fujian Agriculture and Forestry University;UN(China)Center for Fruit Fly Prevention and Treatment;State Key Laboratory of Ecological Pest Control for Fujian and Taiwan Crops;Key Lab of Biopesticide and Chemical Biology,Ministry of Education,Fuzhou,Fujian 350002,China)
出处
《福建农林大学学报（自然科学版）》
CSCD
北大核心
2019年第6期720-726,共7页
Journal of Fujian Agriculture and Forestry University:Natural Science Edition
基金
福建省自然科学基金青年科技人才创新项目(2018J0504)
福建省自然科学基金面上项目(2016J01114)
关键词
膜翅目
茧蜂标本
形态无损
DNA提取试剂盒
PCR
Hymenoptera
Braconidae specimen
non-destructive morphology
DNA extraction kit
PCR
作者简介
夏慧敏(1994-),女.研究方向:农业昆虫与害虫防治.Email:523441201@qq.com;通信作者:陈湜(1984-),女,助理研究员.研究方向:农业昆虫与害虫防治.Email:csalanine@163.com.
