西花蓟马的SCAR分子检测技术

SCAR marker for rapid identification of the western flower thrips,Frankliniella occidentalis (Pergande) (Thysanoptera:Thripidae)

西花蓟马Frankliniella occidentalis(Pergande)是一种世界性入侵害虫,寄主范围广,危害严重,2003年首次在我国发生危害,并有进一步扩散蔓延的趋势。针对蓟马类害虫虫体微小、形态相似,难以准确快速区分的问题,采用特征序列扩增区域(SCAR)标记技术,以西花蓟马及与之同域发生的其他种类蓟马为对象,筛选出1对西花蓟马特异性引物(FOMF/FOMR),其扩增片段大小为320bp。种特异性检测结果显示,该对引物只对西花蓟马的基因组DNA具有扩增能力,对同域发生的花蓟马F.intonsa(Trybom)、禾花蓟马F.tenuicornis(Uzel)、烟蓟马Thripstabaci L.等41种蓟马不具有扩增效果。该对引物不仅对不同虫态的西花蓟马具有扩增能力,而且在西花蓟马发生地的寄主植物组织内亦检测到了其卵的存在。同时,该检测技术灵敏度高,对成虫的最低检出阈值为1/160头。本检测技术在口岸检疫以及花卉、蔬菜和种苗调运中的害虫检测和监测中具有重要意义。

The western flower thirps,Frankliniella occidentalis （Pergande）,is a worldwide pest,causing serious damage to a wide range of host plants.It was first reported in China in 2003,with a trend of further spreading.Most of thrips species are small in size and similar in morphology,which makes them hard to be distinguished quickly.In the present research,a pair of SCAR （sequence characterized amplified regions） primers （FOMF/FOMR） which are specific to western flower thrips was developed by using other nine familiar thrips species as the control.The fragment amplified by the primers FOMF/FOMR was 320 bp in length.Specificity tests performed with this pair of primers showed that all F.occidentalis specimens were detected positively and no cross reactions with other 41 thrips species,including F.intonsa （Trybom）,F.tenuicornis （Uzel） and Thrips tabaci L.,were detected.The method was tested on single male and female adult,single pre-pupa and pupa,single 1st-and 2nd-instar larva,and even single egg,and proved to be applicable for these stages of F.occidentalis.Moreover,the 320 bp genomic DNA fragment was clearly identified in the host plant tissues collected in F.occidentalis infested areas.When the dilution was 1/160 of a whole female adult of F.occidentalis,the 320 bp DNA fragment could be identified for all replicates.The technique should be useful in quarantine at ports and in pest detection and monitoring during transportation of flowers,vegetables and seedlings.