Selection of reference genes for gene expression studies in human neutrophils by real-time PCR

• Published in: BMC molecular biology Vol. 6; no. 1; p. 4
• Main Authors: Zhang, Xiaozhu, Ding, Lily, Sandford, Andrew J
• Format: Journal Article
• Language: English
• Published: England BioMed Central Ltd 18.02.2005; BioMed Central
• Subjects: Actins - analysis; beta 2-Microglobulin - analysis; Gene Expression - genetics; Gene Expression Profiling - standards; Genes; GTP-Binding Proteins - analysis; Humans; Hypoxanthine Phosphoribosyltransferase - analysis; Neoplasm Proteins - analysis; Neutrophils - metabolism; Polymerase Chain Reaction - methods; Polymerase Chain Reaction - standards; Receptors for Activated C Kinase; Receptors, Cell Surface - analysis; Reference Standards; Ribosomal Proteins - analysis; RNA - chemistry; RNA - isolation & purification
• ISSN: 1471-2199; 1471-2199
• DOI: 10.1186/1471-2199-6-4

Reference genes, which are often referred to housekeeping genes, are frequently used to normalize mRNA levels between different samples. However the expression level of these genes may vary among tissues or cells, and may change under certain circumstances. Thus the selection of reference gene(s) is critical for gene expression studies. For this purpose, 10 commonly used housekeeping genes were investigated in isolated human neutrophils. Initial screening of the expression pattern demonstrated that 3 of the 10 genes were expressed at very low levels in neutrophils and were excluded from further analysis. The range of expression stability of the other 7 genes was (from most stable to least stable): GNB2L1 (Guanine nucleotide binding protein, beta polypeptide 2-like 1), HPRT1 (Hypoxanthine phosphoribosyl transferase 1), RPL32 (ribosomal protein L32), ACTB (beta-actin), B2M (beta-2-microglobulin), GAPD (glyceraldehyde-3-phosphate dehydrogenase) and TBP (TATA-binding protein). Relative expression levels of the genes (from high to low) were: B2M, ACTB, GAPD, RPL32, GNB2L1, TBP, and HPRT1. Our data suggest that GNB2L1, HPRT1, RPL32, ACTB, and B2M may be suitable reference genes in gene expression studies of neutrophils.