The normalization of gene expression data in melanoma: investigating the use of glyceraldehyde 3-phosphate dehydrogenase and 18S ribosomal RNA as internal reference genes for quantitative real-time PCR.
We examined a panel of 26 melanoma and fibroblast samples (tissues and cultured cells) to evaluate the suitability of two commonly used housekeeping genes, glyceraldehyde 3-phosphate dehydrogenase (GAPDH) and 18S ribosomal RNA (rRNA), for quantitative real-time PCR. Both genes showed significant variations within the individual cell line and tissue groups. Although no overall trends were observed in the expression of the 18S rRNA, GAPDH was up-regulated in melanoma tissue and cultured cells compared with the corresponding normal samples. In melanoma and fibroblast cell lines and tissues, absolute quantification appears to be more appropriate than normalizing messenger RNA (mRNA) expression via GAPDH or 18S rRNA housekeeping genes.
Department of Chemistry and Biochemistry, Florida Atlantic University, 777 Glades Road, Boca Raton, FL 33431-0991, USA.
Orsolya
O
Giricz
Janelle L
JL
Lauer-Fields
Gregg B
GB
Fields
D045744:Cell Line, Tumor; D020869:Gene Expression Profiling / Q000592:standards*; D015972:Gene Expression Regulation, Neoplastic; D005987:Glyceraldehyde-3-Phosphate Dehydrogenases / Q000235:genetics*; D006801:Humans; D008545:Melanoma / Q000235:genetics*; D016133:Polymerase Chain Reaction / Q000379:methods*; D012333:RNA, Messenger / Q000235:genetics; D012337:RNA, Ribosomal, 18S / Q000235:genetics*; D012015:Reference Standards; D013997:Time Factors
D012333:RNA, Messenger; D012337:RNA, Ribosomal, 18S; D005987:Glyceraldehyde-3-Phosphate Dehydrogenases