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针对粉虱类害虫难以准确快速地进行形态鉴别的问题,以局部发生的黑刺粉虱Aleurocanthus spiniferus(Quaintance)为对象,采用特征序列扩增区域(SCAR)标记法,研究其快速分子检测技术。利用SCAR标记技术获得了长度为987bp的黑刺粉虱特异性片段(GenBank登录号为FJ613323),根据此片段的碱基序列设计黑刺粉虱特异性引物1对(AS-F518/AS-R938),其扩增片段为421bp。种特异性检验结果显示,该对引物只对黑刺粉虱的基因组具有扩增能力,对同域发生的桔绿粉虱Dialeurodes citri(Ashmead)以及其他种类的粉虱如烟粉虱Bemisia tabaci(Gennadius)B型、Q型、ZHJ-1型和ZHJ-2型,温室粉虱Trialeurodes vaporariorum(Westwood)以及螺旋粉虱Aleurodicus disperses(Russell)等的基因组不具有扩增效果。该引物不仅对成虫具有良好的扩增效能,对卵、2龄若虫和拟蛹等亦具有同样的扩增能力,其最低检出限为1/1920头成虫。该技术体系的建立在茶树和柑桔苗木调运的害虫检疫和监测/检测中具有重要意义。
Aiming at the problem that whitefly pests are difficult to identify accurately and rapidly, the rapid detection of the molecular markers of Aleurocanthus spiniferus (Quaintance) was studied by using the sequence of amplified region (SCAR) marker. A specific fragment of GenBank accession No. FJ613323 with a length of 987bp was obtained by SCAR marker. One pair of primers (AS-F518 / AS-R938) was designed based on the nucleotide sequence of this fragment ), The amplified fragment was 421bp. Species-specific test results showed that the pair of primers only had the ability to amplify the genome of B. tabaci Dialeurodes citri (Ashmead) and other species of whiteflies such as Bemisia tabaci Gennadius) genotypes B, Q, ZHJ-1 and ZHJ-2, Trialeurodes vaporariorum (Westwood) and Aleurodicus disperses (Russell) of the greenfly whiteflies did not have any amplification effect. The primer not only has good amplification effect on adults, but also has the same amplification ability on eggs, 2nd instar nymphs and pseudo-pupae, with the minimum detection limit of 1/1920 adult worms. The establishment of this technical system is of great importance in the quarantine and monitoring / detection of pests for the transportation of tea trees and citrus seedlings.