Validation of reference genes for RT-qPCR relative expression analysis during cyst-to-early adult development of...

The development of the zoonotic parasitic tapeworm Taenia solium from larval to adult involves significant but often clinically overlooked events crucial in cestode biology. The early-adult events can be studied in vitro, providing a valuable model to examine scolex evagination, strobilation, and worm development. With some transcriptomes being reported, single-gene relative expression analysis using reverse transcription of RNA (RT) followed by quantitative PCR (qPCR) is valuable to confirm differential expression and study gene regulation during parasite development. However, accurate comparisons with this approach require the validation of endogenous reference genes (RGs). This study identifies stable RGs for normalizing transcript expression data in Taenia. We examined 12 candidate RGs across three "early tapeworm" phases grown in culture. Transcripts were evaluated with RNA-seq and qPCR. Stability rankings were generated using geNorm and NormFinder (for RNA-seq) and RefFinder (for qPCR). Transcripts for rpl13 and ef1α were ranked as the most stable and were tested by using them to normalize the expression of h2b and wnt11a, involved in proliferation and strobilation processes.