Accupower 2x greenstar qpcr mastermix rox dye

Total RNA was isolated with TRIzol reagent (Invitrogen, Carlsbad, CA, USA). The cDNA was synthesized from 4 μg of RNA with M-MLV reverse transcriptase (Bioneer Co., Daejeon, Korea). qRT-PCR was performed with AccuPower 2X Greenstar qPCR MasterMix (-ROX Dye) (Bioneer Co., Daejeon, Korea) on a fluorometric thermal cycler (Corbett Research, Mortlake, NSW, Australia). Primers used in the present study are described in Table S2. Gene expression was calculated according to the comparative 2 ^−ΔΔCT method [34 (link)]. The ΔΔCt value for each sample was determined by calculating the difference between the Ct value of the target gene and the Ct value of the reference gene (β-actin). Values were expressed as fold change of the HC group.

Total RNA was isolated from epididymal WAT or skeletal muscle using Ribo Ex (Geneall Biotechnology Co., Ltd., Daejeon, Korea). The cDNA was synthesized from 4 μg total RNA using M-MLV reverse transcriptase (Bioneer Co., Daejeon, Korea). A fluorometric thermal cycler (Rotor GeneTM 2000; Corbett Research, Mortlake, N.S.W., Australia) and AccuPower 2X Greenstar qPCR Master mix (-ROX Dye) (Bioneer Co.) were utilized for Real Time qPCR analysis, and the data were relatively quantified using the ΔΔC_t method [20 (link)]. Glyceraldehyde 3-phosphate dehydrogenase (GAPDH) was used as a reference gene for normalization and the results were expressed as a fold-difference compared to the HFD group. Primers used in the present study are described in supplementary Table S1.