Screening of reference genes for expression analysis in the study of soldier caste differentiation of Formosan subterranean termite Coptotermes formosanus Shiraki

He Du; Wenjing Wu; Xueyi Huang; Zhiqiang Li

doi:10.7717/peerj.7981

Screening of reference genes for expression analysis in the study of soldier caste differentiation of Formosan subterranean termite Coptotermes formosanus Shiraki

PeerJ. 2019 Nov 5:7:e7981. doi: 10.7717/peerj.7981. eCollection 2019.

Authors

He Du¹, Wenjing Wu¹, Xueyi Huang¹, Zhiqiang Li¹

Affiliation

¹ Guangdong Key Laboratory of Animal Conservation and Resource Utilization, Guangdong Public Laboratory of Wild Animal Conservation and Utilization, Guangdong Institute of Applied Biological Resources, Guangzhou, China.

Abstract

The Formosan subterranean termite, Coptotermes formosanus Shiraki, is a serious pest worldwide. Juvenile hormone analog (and its analogs such as methoprene) can induce the transformation of the worker caste into soldier caste in C. formosanus. However, several factors, such as feeding substrate and colony origin, influence the proportion of solider formation. The molecular mechanism of worker to soldier transformation of C. formosanus is still not clear. RT-qPCR is a powerful tool for molecular studies. Accurate gene quantification by the relative quantification method requires a stable expressed gene as the reference gene. However, no reference genes were available for this species in the methoprene bioassay. To study the problem of gene response to methoprene by RT-qPCR we have to first screen reference genes in C. formosanus. Workers were fed with methoprene. Termites were collected during the methoprene bioassay and separated into head and thorax+abdomen. Expression profiles of 10 candidate reference genes in the two body part types were investigated using RT-qPCR. The results were analyzed by a set of established methods (geNorm, NormFinder, BestKeeper, and RefFinder) as well as comparative ΔCt method. Our results suggest that RPS18 is the most stably-expressed gene both in the head and thorax+abdomen for expression analysis in the methoprene bioassay of C. formosanus. The screening of suitable reference genes in C. formosanus establishes the foundation for the molecular study of soldier caste differentiation in this species.

Keywords: Expression stability; Methoprene bioassay; Normalization.

Grants and funding

This research was funded by GDAS’ Project of Science and Technology Development (Grant No. 2017GDASCX-0836), Startup Funding of Guangdong Institute of Applied Biological Resources (Grant No. GIABR-gjrc201601), the GDAS Special Project of Science and Technology Development (2018GDASCX-0107), and the National Natural Science Foundation of China (Grant No. 31702068). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.