A comparative assessment of reference genes in mouse brown adipocyte differentiation and thermogenesis in vitro
Trang Huyen Lai,
Jin Seok Hwang,
Quang Nhat Ngo,
Dong-Kun Lee,
Hyun Joon Kim,
Deok Ryong Kim
Affiliations
Trang Huyen Lai
Department of Biochemistry and Convergence Medical Sciences and Institute of Medical Science, Gyeongsang National University, College of Medicine, Jinju, South Korea
Jin Seok Hwang
Department of Biochemistry and Convergence Medical Sciences and Institute of Medical Science, Gyeongsang National University, College of Medicine, Jinju, South Korea
Quang Nhat Ngo
Department of Biochemistry and Convergence Medical Sciences and Institute of Medical Science, Gyeongsang National University, College of Medicine, Jinju, South Korea
Dong-Kun Lee
Department of Physiology and Convergence Medical Sciences and Institute of Medical Science, Gyeongsang National University, College of Medicine, Jinju, South Korea
Hyun Joon Kim
Department of Anatomy and Convergence Medical Sciences and Institute of Medical Science, Gyeongsang National University, College of Medicine, Jinju, South Korea
Deok Ryong Kim
Department of Biochemistry and Convergence Medical Sciences and Institute of Medical Science, Gyeongsang National University, College of Medicine, Jinju, South Korea
ABSTRACTAdipogenic differentiation and thermogenesis in brown adipose tissue (BAT) undergo dynamic processes, altering phenotypes and gene expressions. Proper reference genes in gene expression analysis are crucial to mitigate experimental variances and ensure PCR efficacy. Unreliable reference genes can lead to erroneous gene expression quantification, resulting in data misinterpretation. This study focused on identifying suitable reference genes for mouse brown adipocyte research, utilizing brown adipocytes from the Ucp1-luciferase ThermoMouse model. Comparative analysis of gene expression data under adipogenesis and thermogenesis conditions was conducted, validating 13 housekeeping genes through various algorithms, including DeltaCq, BestKeeper, geNorm, Normfinder, and RefFinder. Tbp and Rer1 emerged as optimal references for Ucp1 and Pparg expression in brown adipogenesis, while Tbp and Ubc were ideal for the expression analysis of these target genes in thermogenesis. Conversely, certain conventional references, including Actb, Tubb5, and Gapdh, proved unstable as reference genes under both conditions. These findings stress the critical consideration of reference gene selection in gene expression analysis within specific biological systems to ensure accurate conclusions.
