TY - JOUR
T1 - Evaluation of reference genes for normalization of quantitative real time PCR in non-small cell lung cancer
AU - Endo, Chiaki
AU - Sun, Zhifu
AU - Molina, Julian R.
AU - Attewell, John R.
AU - Yang, Ping
PY - 2007/1/1
Y1 - 2007/1/1
N2 - To compare gene expression levels of lung cancer by real time quantitative reverse transcription polymerase chain reaction (qRT-PCR), choosing appropriate control genes for normalization of RNA input is important. We examined whether frequently used control genes are really appropriate. Eighteen lung cancer tissue samples were used in this study. Gene expression levels were measured by qRT-PCR and compared to the data generated by a DNA microarray experiment. The expression levels of four housekeeping genes that are most frequently used as control genes for qRT-PCR published reports in lung cancer were exammed. Four test genes (TP53, laminin γ2, TGFα and cathepsin L2) were also examined. These genes had significantly higher expression in the high aggressive disease group. When taken as single genes, the expression levels of these so called control genes (glyceraldehydes-3phosphate dehydrogenase, TATA box binding protein, β actin and β2 microgloblin) were not consistent among the tested samples. However, the geometric mean of the threshold cycle of the TATA box binding protein and β2 microglobulin showed less variation than that of the single control gene. These results indicate that normalization by multiple control genes is more appropriate for comparison of gene expression levels of lung cancer when using qRT-PCR analysis.
AB - To compare gene expression levels of lung cancer by real time quantitative reverse transcription polymerase chain reaction (qRT-PCR), choosing appropriate control genes for normalization of RNA input is important. We examined whether frequently used control genes are really appropriate. Eighteen lung cancer tissue samples were used in this study. Gene expression levels were measured by qRT-PCR and compared to the data generated by a DNA microarray experiment. The expression levels of four housekeeping genes that are most frequently used as control genes for qRT-PCR published reports in lung cancer were exammed. Four test genes (TP53, laminin γ2, TGFα and cathepsin L2) were also examined. These genes had significantly higher expression in the high aggressive disease group. When taken as single genes, the expression levels of these so called control genes (glyceraldehydes-3phosphate dehydrogenase, TATA box binding protein, β actin and β2 microgloblin) were not consistent among the tested samples. However, the geometric mean of the threshold cycle of the TATA box binding protein and β2 microglobulin showed less variation than that of the single control gene. These results indicate that normalization by multiple control genes is more appropriate for comparison of gene expression levels of lung cancer when using qRT-PCR analysis.
KW - Control gene
KW - Non-small cell lung cancer
KW - Normalization
KW - RNA
KW - Real time PCR
KW - Squamous cell carcinoma
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U2 - 10.3923/ijcr.2007.1.12
DO - 10.3923/ijcr.2007.1.12
M3 - Article
AN - SCOPUS:34249294491
VL - 3
SP - 1
EP - 12
JO - International Journal of Cancer Research
JF - International Journal of Cancer Research
SN - 1811-9727
IS - 1
ER -