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While these results demonstrate considerable consistency between induction measured by short-read sequencing of oligo dT) primed, double-stranded cDNA and induction measured by traditional microarray methods, they also suggest that higher depth of coverage may be needed to accurately assay weakly induced genes.
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Candidates were confirmed by discriminating homozygote and heterozygote populations using a fluorescence solution method and two colorimetric microarray methods on polycarbonate chips.
Comparing with microarray methods, deep RNA sequencing (RNA-Seq) has been more widely used in different cell biology processes due to advantages of unbias, high-throughput, and sensitivity.
Expanded microarray methods are provided in the Detailed Materials and Methods (Appendix S1).
Details of RNA extraction and microarray methods have been described elsewhere [21], [22].
For gene expression data, we used the normalized log ratios of Cy5/Cy3 (see Microarray Methods).
The Robust Multi-Array Average (RMA) algorithm was used and data was also analyzed by the significant analysis of microarray methods (SAM).
GL26nu, GL26B6 and GL26B6V cells (recovered as in microarray methods above) were cultured at 100,000 per well in 1 mL complete RPMI.
Microarray methods were modified from [ 92].
Detailed DNA microarray methods and microarray data associated with this publication are available through the Gene Expression Omnibus [GEO GSE29801].
Sample preparation and microarray methods followed those of our previous study [ 7].
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