Real-Time PCR Analysis of Metabolism-Related Genes in a Long-Lived Model of C. elegans.

Abstract

In the nematode Caenorhabditis elegans, the mammalian tumor suppressor p53 ortholog CEP-1(C. elegans p53-like protein) is associated not only with the stress response, germline apoptosis, and meiotic chromosome segregation but also with longevity through the modification of energy metabolism during aging. The mitochondrial respiration-related gene sco-1 in C. elegans is orthologous to the human SCO1 gene and a target of p53/CEP-1. Using quantitative real-time polymerase chain reaction (PCR) analysis, we recently found that the expression levels of sco-1 gene were increased in wild-type C. elegans in an aging-related manner and decreased in long-lived cep-1 mutants. Here, we describe the relative quantitative strategy using a commercial real-time PCR system to detect more accurately differences in the levels of expressed genes between long-lived and wild-type C. elegans strains. To estimate the expression levels of target genes compared with wild-type using relative quantification, we used the expression levels of an endogenous control gene, such as a housekeeping gene. In addition, it is critical to normalize differences in the expression levels of the common housekeeping gene among the strains analyzed for an accurate comparison of the quantitative expression levels of target genes.