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Differential expression analysis was done for microarray expression intensities of 679 bovine miRNAs with SAM (significance analysis of microarrays) method at delta parameter of 0.6 and q FDR < 12%.
Microarray expression intensities were preprocessed using the 'GCRMA' (v2.14.1) [ 38] package in Bioconductor, which corrected backgrounds based on calculated affinities of probe sequences, quantile-quantile normalized intensities, and summarized expressions of probe sets through median-polish and log2 transformed expression values.
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However, we performed our other analyses with the genes providing Microarray expression intensity and the number of genes (in Microarray expression dataset) for GFPψ genes and GLψ genes are 1443 and 10192 respectively (Additional file 2).
For microarray data, expression intensities for each probeset were computed as MAS5 signal intensities, and subsequently quantile-normalized and transformed to homogeneous variance using the "S10" transform available in the MSCL Analyst's Toolbox (http://abs.cit.nih.gov/MSCLtoolbox/).nih.gov/MSCLtoolbox/
For microarray data, we retrieved the expression intensities of each probe set from NCBI's GEO database.
The microarray reported gene expression intensity is an RMA signal in log2 scale.
5 The rank product statistic ranks genes according to expression intensities within each microarray and calculates the product of these ranks across multiple microarrays.
Using a DNA microarray, we measured and compared the expression intensities of all 4345 genes of each strain cultured under three different conditions.
A selection of genes with high expression intensities identified by microarray were validated by quantitative Real-Time PCR (Additional file 2: Supplementary Methods; Additional file 2: Figure S4).
Microarray expression profiles are extracted from signal intensities reflecting the amount of hybridized mRNA to spotted DNA whereas the above mentioned sequencing-based technologies provide expression levels that are absolute values computed as the number of transcripts observed for individual genes.
A number of algorithms are available for calculating the expression intensities on Affymetrix microarrays.
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