Taurine Biosynthesis Gene Expression in BSF

Quantitative PCR (qPCR) was used to evaluate the mRNA levels of the putative genes associated with the taurine biosynthesis in BSF larvae and prepupae. Transcript levels of ado, cdo, csad, and gad were quantified using the QuantiTect SYBR^®Green PCR Kit (Qiagen), 1:20 diluted cDNA samples, and gene-specific qPCR primers (Table S2) with a Rotor-Gene Q 2 plex Hrm thermocycler (Qiagen). The amplification efficiency and the specificity of each reaction were evaluated as reported elsewhere [37 (link)]. For each reaction, six biological replicates were run in duplicate together with minus reverse transcriptase and no template controls. Raw data of target genes were corrected using the normalization factor calculated by the GeNorm Software from three suitable reference genes, elongation factor (ef1-α), 18s ribosomal RNA (18s rRNA), and 16s ribosomal RNA (16s rRNA), as described by [38 (link)].