Short Communication: Optimization of semi-quantitative RT PCR analysis for CPT I gene expression in Rainbow trout (Oncorhynchus mykiss)

Abstract

A key enzyme in mitochondrial βoxidation, carnitine palmitoyltransferase (CPT) I, is transcriptionally regulated in mammals, but this enzyme also experiences allosteric modulations (Harano et al., 1985; Murthy and Pande, 1987; Bezaire et al., 2004). CPT I is located on the inner side of the outer mitochondrial membrane and catalyses the conversion of acyl-CoA to fatty acylcarnitine (Kerner and Hoppel, 2000; Price et al., 2000). Quantitative RT-PCR is a reliable technique for measuring transcripts in small amounts of tissue (Spriewald et al., 2000). With this technique, multiple mRNAs can be assayed simultaneously in a relatively short period of time. Here we describe the standard procedure, optimized in our laboratory, to assess CPT I levels with βactin as an internal control in rainbow trout, and all the necessary controls to ensure a quantitative analysis.