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Open Access Highly Accessed Methodology article

A rapid and inexpensive labeling method for microarray gene expression analysis

Mario Ouellet12, Paul D Adams124, Jay D Keasling1234 and Aindrila Mukhopadhyay12*

Author Affiliations

1 The Joint Bioenergy Institute, Lawrence Berkeley National Laboratory, Emeryville, USA

2 Physical Biosciences Division, Lawrence Berkeley National Laboratory, Berkeley, USA

3 Department of Chemical Engineering, University of California, Berkeley, USA

4 Department of Bioengineering, University of California, Berkeley, USA

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BMC Biotechnology 2009, 9:97  doi:10.1186/1472-6750-9-97

Published: 25 November 2009

Abstract

Background

Global gene expression profiling by DNA microarrays is an invaluable tool in biological research. However, existing labeling methods are time consuming and costly and therefore often limit the scale of microarray experiments and sample throughput. Here we introduce a new, fast, inexpensive method for direct random-primed fluorescent labeling of eukaryotic cDNA for gene expression analysis and compare the results obtained on the NimbleGen microarray platform with two other widely-used labeling methods, namely the NimbleGen-recommended double-stranded cDNA protocol and the indirect (aminoallyl) method.

Results

Two total RNA samples were labeled with each method and hybridized to NimbleGen expression arrays. Although all methods tested here provided similar global results and biological conclusions, the new direct random-primed cDNA labeling method provided slightly better correlation between replicates compared to the other methods and thus increased ability to find statistically significant differentially expressed genes.

Conclusion

The new direct random-primed cDNA labeling method introduced here is suitable for gene expression microarrays and provides a rapid, inexpensive alternative to existing methods. Using NimbleGen microarrays, the method produced excellent results comparable to those obtained with other methods. However, the simplicity and cost-effectiveness of the new method allows for increased sample throughput in microarray experiments and makes the process amenable to automation with a relatively simple liquid handling system.