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07:04
Performing Custom MicroRNA Microarray Experiments
Published on: October 28, 2011
Optimization of cDNA microarrays procedures using criteria that do not rely on external standards
Torunn Bruland1, Endre Anderssen, Berit Doseth
1Department of Cancer Research and Molecular Medicine, Faculty of Medicine, Norwegian University of Science and Technology (NTNU), N-7489 Trondheim, Norway. torunn.bruland@ntnu.no
BMC Genomics
|October 24, 2007
Summary
Optimizing microarray gene expression analysis is challenging without external standards. A new high-contrast versus self-self method (HCSSM) improves gene identification using internal controls and minimal arrays.
Area of Science:
- Molecular Biology
- Genomics
- Bioinformatics
Background:
- Microarray gene expression measurement is complex with many variables.
- Lack of standard calibration samples hinders evaluation of procedural improvements.
- Optimizing microarray procedures requires methods independent of external standards.
Purpose of the Study:
- To optimize laboratory and data processing steps in microarray analysis.
- To develop a method for evaluating procedural improvements without external standards.
- To maximize the identification of differentially expressed genes.
Main Methods:
- Performed cDNA microarray experiments with high-contrast and self-self hybridizations.
- Developed a high-contrast versus self-self method (HCSSM) using internal controls.
- Investigated effects of blocking reagent dose, filtering, and background correction on gene identification.
- Utilized false discovery rate (FDR) estimation based on a null distribution from self-self experiments.
Main Results:
- The high-contrast versus self-self method (HCSSM) requires only four microarrays.
- Optimal conditions included 250 ng LNA dT blocker, no background correction, and weight-based filtering.
- Background correction method choice significantly impacted the number of differentially expressed genes.
- Cross-platform validation confirmed the real increase in identified differentially expressed genes.
Conclusions:
- HCSSM offers a simple and effective approach for optimizing microarray procedures without external standards.
- The method is applicable to both long oligo-probe and cDNA microarrays.
- This optimization strategy is valuable for various organisms, including those with incomplete genome information.
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