%0 Journal Article
%T Evaluation of suitable reference genes for normalization of real-time reverse transcription PCR analysis in colon cancer
%A Lise S£¿rby
%A Solveig N Andersen
%A Ida RK Bukholm
%A Morten B Jacobsen
%J Journal of Experimental & Clinical Cancer Research
%D 2010
%I BioMed Central
%R 10.1186/1756-9966-29-144
%X In this study, expression of 16 commonly used reference genes was quantified in tumour tissue and individual-matched normal mucosa in 18 non-metastatic colon cancer patients and 20 colon cancer patients with distant metastases using TaqMan Low Density Array (TLDA). The expression stability was determined and compared by means of geNorm and NormFinder.Two pairs of genes, HPRT1/PPIA and IPO8/PPIA, were identified to be suitable to normalize gene expression data in metastatic and non-metastatic colon cancer patients, according to geNorm and NormFinder respectively.We propose a standardized approach of finding the most suitable reference gene(s) in every qRT-PCR experiment using TLDA.qRT-PCR is one of the most sensitive methods for mRNA detection and quantification. The method has also become the preferred method for validating results obtained by other techniques, such as microarray [1]. There are differences among different qRT-PCR assays due to biological and technical variations [2,3]. In order to identify truly gene specific variations it is important to use a suitable normalization method. One of the most commonly used approaches involves relative quantification of target genes against one or more reference genes which are thought to be stably expressed in the examined tissue [4]. There have been a number of reports that demonstrate that the expression levels of putative reference genes vary extensively in different tissues and diseases and thus are unsuitable for normalization purposes [5-15]. Consequently, each research group has to validate multiple reference genes in their own experimental setup and normalize qRT-PCR data against a few reference genes tested from independent pathways using at least one algorithm. It appears that improvements in methods of identifying reference genes are more important than the identification of the particular reference genes themselves [16].It has been argued for use of multiple genes in the normalization of qRT-PCR analysis a
%U http://www.jeccr.com/content/29/1/144