Evaluation of reference genes for expression studies in the broad mite, Polyphagotarsonemus latus (Acari: Tarsonemidae)

Abstract

The broad mite, Polyphagotarsonemus latus (Banks) (Acari: Tarsonemidae), has been a devastating pest of several important agricultural and horticultural crops. Studies on the molecular mechanisms of adaptation to extrinsic conditions, e.g. acaricide exposure and temperature stress could aid in revealing the mite’s xenobiotic metabolism and quick adaptation to rapidly changing environments, respectively. This is best studied by the gene expression patterns, which could be accomplished through real-time quantitative reverse transcription PCR (qRT-PCR). Since real-time qRT-PCR studies require the selection of one or more reference genes whose expression patterns might vary across treatments, it is indispensable to validate their expression stability. In this study, the expression stability of six candidate reference genes viz., actin, elongation factor (EF-1α), glyceraldehyde-3-phosphate-dehydrogenase (GAPDH), ribosomal protein 49 (RP49), ribosomal protein S18 (RPS18) and β-tubulin (TUB) was investigated through real-time qRT-PCR experiments. By integrating the results of geNorm, NormFinder, BestKeeper, comparative ΔCt and RefFinder algorithms, RPS18/RP49 and EF-1α/RPS18 were identified as highly suitable reference genes for acaricide treatment and temperature stress treatments, respectively. RPS18 was recognized as the most suitable reference gene for both treatments which will ensure the accuracy of target gene expressions in studies related to xenobiotic metabolism and stress tolerance in P. latus.