Screening and Identification of Reference Genes for Paracarophenax alternatus

In this study, we focused on the screening and identification of reference genes for Paracarophenax alternatus Xu and Zhang. The laboratory population was used as the laboratory population, and samples were collected from mites at four different stages, including physogastry, viviparous, 5 d viviparous and phoresy. Then, the expression levels of seven candidate reference genes (α-tubulin, β-tubulin, RPS18, RPL13, GAPDH, EF1A, SDHA) were detected through qRT-PCR. Melting curves showed good gene specificity, and the amplification efficiency ranged from 90% to 102%. ΔCt analysis indicated that GAPDH was the most stable reference gene. The GeNorm software determined that the optimal number of reference genes was two, with GAPDH and RPS18 forming the most stable combination, and NormFinder identified RPS18 as the most stable reference gene. Although the BestKeeper software suggested that EF1A was the most stable, its p-value exceeded 0.05, rendering it unsuitable for use as a reference gene. Finally, through the RefFinder network tool, the most stable reference genes were identified as GAPDH and RPS18.