基于种特异性COI标记的花生蚜快速鉴定技术

Rapid Identification of Aphis craccivora(Homoptera: Aphididae) Based on Species-specific COI Marker

针对花生蚜虫体型小、在田间不易准确快速识别的问题。本文采用基于线粒体DNA细胞色素C氧化酶亚基I(mitochondrial DNA cytochrome coxidase subunit I,mtDNA COI)基因的种特异(species-specific COI)PCR方法,研究了其快速鉴定的分子技术。研究利用mtDNA COI基因通用型引物F-LCO-1490/R-HCO-700ME获得青岛地区花生蚜虫COI基因序列,并根据测序结果设计六对花生蚜特异引物,同时以同一花生田采集的、与花生蚜同生境节肢动物、线虫、软体动物的166个DNA样本为模板对六对引物的种特异性进行了检测。PCR结果显示,引物AC_F2/AC_R2对花生蚜基因组模板有特异扩增能力,片段长度为299bp;且该对引物在与花生蚜同生境节肢动物、线虫、软体动物样本中均未扩增出条带,显示出此种特异性引物对其他物种不具有交叉反应扩增能力,可用于花生蚜的快速检测及预测预报,同时对本地天敌对花生蚜捕食作用的检测也具有重要意义。

Morphological identification of Aphis craccivora is limited because of small size,and it is difficult to be accurately and quickly identified in the field.In this paper,a species-specific COI PCR method based on mitochondrial DNA cytochrome coxidase subunit I(mtDNA COI)gene was used to the rapid identification of peanut aphid.COI genesequence of peanut aphid in Qingdao region was obtained by mtDNA universal primers COI gene F-LCO-1490/R-HCO-700ME,and six specific primers were designed according to the sequence.The primer AC_F2/AC_R2 had specific amplification ability to peanut aphid(299bp),and exhibited specificity for peanut aphid when screened for cross-reactivity against 166 nontarget DNA extractions(88 nontarget taxa).The results showed that the primer AC_F2/AC_R2 could be used in the rapid identification,early predation and predator evaluation of peanut aphid.