Wosula Everlyne N, Chen Wenbo, Amour Massoud, Fei Zhangjun, Legg James P
International Institute of Tropical Agriculture, P.O. Box 34441 Dar es Salaa, Tanzania.
Boyce Thompson Institute, 533 Tower Rd, Ithaca, NY 14853, USA.
Insects. 2020 May 14;11(5):305. doi: 10.3390/insects11050305.
is a cryptic species complex that requires the use of molecular tools for identification. The most widely used approach for achieving this is the partial sequencing of the mitochondrial DNA cytochrome oxidase I gene (). A more reliable single nucleotide polymorphism (SNP)-based genotyping approach, using Nextera restriction-site-associated DNA (NextRAD) sequencing, has demonstrated the existence of six major haplogroups of on cassava in Africa. However, NextRAD sequencing is costly and time-consuming. We, therefore, developed a cheaper and more rapid diagnostic using the Kompetitive Allele-Specific PCR (KASP) approach. Seven sets of primers were designed to distinguish the six haplogroups based on the NextRAD data. Out of the 152 whitefly samples that were tested using these primer sets, 151 (99.3%) produced genotyping results consistent with NextRAD. The KASP assay was designed using NextRAD data on whiteflies from cassava in 18 countries across sub-Saharan Africa. This assay can, therefore, be routinely used to rapidly diagnose cassava by laboratories that are researching or monitoring this pest in Africa. This is the first study to develop an SNP-based assay to distinguish whiteflies on cassava in Africa, and the first application of the KASP technique for insect identification.
是一个隐秘的物种复合体，需要使用分子工具进行鉴定。实现这一目标最广泛使用的方法是对线粒体DNA细胞色素氧化酶I基因（）进行部分测序。一种更可靠的基于单核苷酸多态性（SNP）的基因分型方法，即使用Nextera限制性位点相关DNA（NextRAD）测序，已经证明在非洲木薯上存在六个主要的单倍群。然而，NextRAD测序成本高且耗时。因此，我们开发了一种使用竞争性等位基因特异性PCR（KASP）方法的更便宜、更快速的诊断方法。根据NextRAD数据设计了七组引物，以区分六个单倍群。在使用这些引物组测试的152个粉虱样本中，151个（99.3%）产生了与NextRAD一致的基因分型结果。KASP分析是根据撒哈拉以南非洲18个国家木薯粉虱的NextRAD数据设计的。因此，该分析可由在非洲研究或监测这种害虫的实验室常规用于快速诊断木薯粉虱。这是第一项开发基于SNP的分析方法来区分非洲木薯上的粉虱的研究，也是KASP技术在昆虫鉴定中的首次应用。
