Laboratory of Molecular Entomology and Bee Pathology, Ghent University, Krijgslaan 281, B-9000, Ghent, Belgium.
Animal Genetics Laboratory, Ghent University, Heidestraat 19, B-9820, Merelbeke, Belgium.
BMC Vet Res. 2021 Apr 30;17(1):179. doi: 10.1186/s12917-021-02886-x.
The varroa mite is one of the main causes of honey bee mortality. An important mechanism by which honey bees increase their resistance against this mite is the expression of suppressed mite reproduction. This trait describes the physiological inability of mites to produce viable offspring and was found associated with eight genomic variants in previous research.
This paper presents the development and validation of high-throughput qPCR assays with dual-labeled probes for discriminating these eight single-nucleotide variants. Amplicon sequences used for assay validation revealed additional variants in the primer/probe binding sites in four out of the eight assays. As for two of these the additional variants interfered with the genotyping outcome supplementary primers and/or probes were developed. Inclusion of these primers and probes in the assay mixes allowed for the correct genotyping of all eight variants of interest within our bee population.
These outcomes underline the importance of checking for interfering variants in designing qPCR assays. Ultimately, the availability of this assay allows genotyping for the suppressed mite reproduction trait and paves the way for marker assisted selection in breeding programs.
瓦螨是导致蜜蜂死亡的主要原因之一。蜜蜂增强其对这种螨虫的抵抗力的一个重要机制是抑制螨虫繁殖的表达。这一特性描述了螨虫无法产生有活力后代的生理能力，并且在之前的研究中发现与八个基因组变体有关。
本文介绍了使用双标记探针开发和验证高通量 qPCR 检测方法，以区分这八个单核苷酸变体。用于检测验证的扩增子序列在八个检测方法中的四个中在引物/探针结合位点中发现了其他变体。对于其中两个，额外的变体干扰了基因分型结果，因此开发了补充引物和/或探针。在检测混合物中包含这些引物和探针，使得可以在我们的蜜蜂群体中正确地对所有八个感兴趣的变体进行基因分型。
这些结果强调了在设计 qPCR 检测方法时检查干扰变体的重要性。最终，该检测方法的可用性允许对抑制的螨虫繁殖特性进行基因分型，并为在繁殖计划中进行标记辅助选择铺平了道路。
