Abstract
Real-time quantitative PCR (RT-qPCR) is the most advanced and commonly used method for quantification of gene expression levels because of its high sensitivity, specificity and accuracy. The analytical capability of RT-qPCR, on the other hand, is dependent on precise normalization by using the most stable reference genes. The present study was undertaken to find the best-suited reference gene for normalization of qPCR data for ontogenetic studies of striped murrel. Four commonly used and constitutively expressed genes, namely beta-actin (β-actin), glyceraldehyde-3-phosphate dehydrogenase (GAPDH), elongation factor 1-alpha (EF1α) and 18 S ribosomal protein (18 S RNA) were screened and evaluated for their expression stability in developing larvae from 1 to 35 days post hatch (dph). The stability and suitability of these genes were determined by using four different algorithms; namely, delta Ct, BestKeeper, geNorm and NormFinder. The four algorithms together generated a comprehensive ranking EF1α being the most stable, followed by GAPDH and β-actin as the least stable gene. This is the first report evaluating the most suitable reference gene for mRNA expression studies in C. striata, in the context of ontogenetic development.
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