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Gene expression levels assessed by oligonucleotide microarray analysis and quantitative real-time RT-PCR -- how well
Peter B Dallas1, Nicholas G Gottardo, Martin J Firth
1Division of Children's Leukaemia and Cancer Research, Telethon Institute for Child Health Research and Centre for Child Health Research, The University of Western Australia, Perth, Australia. peterd@ichr.uwa.edu.au
BMC Genomics
|April 28, 2005
Summary
Quantitative real-time RT-PCR (qRT-PCR) and microarray gene expression analysis show strong correlation when targeting identical transcripts. However, careful interpretation is needed as some genes show poor correlation.
Area of Science:
- Molecular Biology
- Genomics
- Biotechnology
Background:
- Microarray technology is widely used for gene expression profiling.
- Validating microarray data with independent methods like quantitative real-time RT-PCR (qRT-PCR) is crucial.
- Standardized validation methods are needed for inter-laboratory data comparison.
Purpose of the Study:
- To assess the correlation between gene expression levels measured by oligonucleotide microarrays and qRT-PCR.
- To evaluate the reliability of microarray data validation using qRT-PCR.
Main Methods:
- Oligonucleotide microarrays were used to obtain expression scores for 48 human genes.
- Quantitative real-time RT-PCR (qRT-PCR) was employed to measure expression levels of the same genes.
- Microarray data were processed using Robust Multi-array Analysis (RMA) and MAS 5.0 algorithms.
Main Results:
- Strong correlations (r = 0.89) were observed between qRT-PCR and microarray data when targeting identical transcripts.
- Poor correlations were found for 13% of genes with RMA and 16% with MAS 5.0 normalized microarray data.
- Identical transcript targeting is key for reliable correlation between the two methods.
Conclusions:
- Oligonucleotide microarrays and qRT-PCR are complementary technologies for validating gene expression measurements.
- Caution is advised when interpreting gene expression data due to potential discrepancies.
- Methodological choices in data processing can impact the correlation between microarray and qRT-PCR results.