蓟马基因组DNA提取方法的改进
Method improvement for extraction genomic DNA from thrips
张利娟1, 2，沈登荣1, 2，张宏瑞1,2**，张宏伟3，李正跃1, 2
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作者单位：1云南农业大学农业生物多样性与病虫害控制教育部重点实验室昆明650201;2云南农业大学植物保护学院昆明650201；3普洱市宁洱县农业技术推广中心普洱665100
中文关键词：蓟马， DNA提取， PCR
英文关键词：thrips, DNA extraction, PCR
中文摘要： 在昆虫分子生物学的研究中，从昆虫样品中有效地获得总DNA是分子实验成功的前提。但是，常规提取方法由于不能保留昆虫所有的形态特征，这对于体形较小的珍稀标本是不适用的。文中通过对改进的盐析法和STE法与KAc法的对比，发现盐析法和STE法提取的DNA质量明显优于KAc法，并且能够通过针刺从单头蓟马中成功提取DNA而不影响形态鉴定。2种提取方法的优点是单头蓟马在提取过DNA以后，虫体仍然可用以做成永久玻片进行形态鉴定。提取的DNA经实验证明，可以顺利的进行mtDNA-COI和rDNA-ITS2基因序列引物的扩增。
英文摘要： To isolate total DNA from insect samples is a crucial prerequisite for successful PCR reactions. However, regular methods for isolating insect DNA can’t resolve all the morphological characters of insects. Because these methods destroy insects’ morphological characteristics they are obviously not suitable for scarce and small insects such as thrips. In this article, two methods of saltingout and the STE method of extracting genome DNA from individual thrips are compared to KAc. The results suggest that，the quality of DNA samples extracted by the saltingout and STE methods was obviously better and more suitable for PCR than those obtained by the KAc method. Genome DNA was extracted from individual thrips by piercing one side of the specimen’s abdomen with a minute, sterilized pin, thereby avoiding destroying their morphological features. Experiments demonstrated that the extracted DNA was suitable for PCR amplification using the sequencing primers Mitochondrial Cytochrome Oxidase I and the ITS2 region of ribosomal DNA.