Evaluation and normalization of a set of reliable reference genes for quantitative gene expression analysis in -focused cancer research.

Multiple signaling pathways have been discovered to play a role in aging and longevity, including the insulin/IGF-1 signaling system, AMPK pathway, TOR signaling, JNK pathway, and germline signaling. Mammalian serum and glucocorticoid-inducible kinase 1 (), which has been associated with various disorders including hypertension, obesity, and tumor growth, limits survival in by reducing DAF-16/FoxO activity while suppressing FoxO3 activity in human cell culture. provides significant protection for a number of genes associated with human cancer. The best known of these are the (mammalian ortholog ) and CEP-1 (mammalian ortholog ) genes. Therefore, in this study, we aimed to investigate the expression analyzes of , which is overexpressed in many types of mammalian cancer, in mutant lin-35 and to demonstrate the validation of reference genes in wild-type N2 and mutant lin-35 for -focused cancer research. To develop functional genomic studies in , we evaluated the expression stability of five candidate reference genes (, ) by quantitative real-time PCR using five algorithms (geNorm, NormFinder, Delta Ct method, BestKeeper, RefFinder) in N2 and lin-35 worms. According to our findings, and were effective in accurately normalizing the levels of gene expression in N2 and lin-35. and also displayed the most consistent expression patterns, therefore they were utilized to standardize expression level of . Furthermore, our results clearly showed that was upregulated in lin-35 worms compared to N2 worms. Our results highlight the importance of definitive validation using mostly expressed reference genes.