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Selection and validation of endogenous reference genes using a high throughput approach.
- Source :
-
BMC genomics [BMC Genomics] 2004 Aug 13; Vol. 5 (1), pp. 55. Date of Electronic Publication: 2004 Aug 13. - Publication Year :
- 2004
-
Abstract
- Background: Endogenous reference genes are commonly used to normalize expression levels of other genes with the assumption that the expression of the former is constant in different tissues and in different physiopathological conditions. Whether this assumption is correct it is, however, still matter of debate. In this study, we searched for stably expressed genes in 384 cDNA array hybridization experiments encompassing different tissues and cell lines.<br />Results: Several genes were identified whose expression was highly stable across all samples studied. The usefulness of 8 genes among them was tested by normalizing the relative gene expression against test genes whose expression pattern was known. The range of accuracy of individual endogenous reference genes was wide whereas consistent information could be obtained when information pooled from different endogenous reference genes was used.<br />Conclusions: This study suggests that even when the most stably expressed genes in array experiments are used as endogenous reference, significant variation in test gene expression estimates may occur and the best normalization is achieved when data from several endogenous reference genes are pooled together to minimize minimal but significant variation among samples. We are presently optimizing strategies for the preparation of endogenous reference gene mixtures that could yield information comparable to that of data pooled from individual endogenous reference gene normalizations.
- Subjects :
- Carcinoma genetics
Carcinoma pathology
Cell Line
Cell Line, Tumor
DNA, Complementary genetics
Gene Expression Profiling standards
Humans
Melanocytes metabolism
Melanoma genetics
Melanoma pathology
Melanoma secondary
Nevus genetics
Nevus pathology
Nucleic Acid Hybridization
Oligonucleotide Array Sequence Analysis
Organ Specificity
RNA, Messenger genetics
RNA, Neoplasm genetics
Reference Standards
Reproducibility of Results
Reverse Transcriptase Polymerase Chain Reaction
Skin Neoplasms genetics
Skin Neoplasms pathology
Gene Expression Profiling methods
Gene Expression Regulation
Genes
Subjects
Details
- Language :
- English
- ISSN :
- 1471-2164
- Volume :
- 5
- Issue :
- 1
- Database :
- MEDLINE
- Journal :
- BMC genomics
- Publication Type :
- Academic Journal
- Accession number :
- 15310404
- Full Text :
- https://doi.org/10.1186/1471-2164-5-55