title: Impact of normalization on miRNA microarray expression profiling.
abstract: Profiling miRNA levels in cells with miRNA microarrays is becoming a widely used technique. Although normalization methods for mRNA gene expression arrays are well established, miRNA array normalization has so far not been investigated in detail. In this study we investigate the impact of normalization on data generated with the Agilent miRNA array platform. We have developed a method to select nonchanging miRNAs (invariants) and use them to compute linear regression normalization coefficients or variance stabilizing normalization (VSN) parameters. We compared the invariants normalization to normalization by scaling, quantile, and VSN with default parameters as well as to no normalization using samples with strong differential expression of miRNAs (heart-brain comparison) and samples where only a few miRNAs are affected (by p53 overexpression in squamous carcinoma cells versus control). All normalization methods performed better than no normalization. Normalization procedures based on the set of invariants and quantile were the most robust over all experimental conditions tested. Our method of invariant selection and normalization is not limited to Agilent miRNA arrays and can be applied to other data sets including those from one color miRNA microarray platforms, focused gene expression arrays, and gene expression analysis using quantitative PCR.
authors.affiliation: Lausanne DNA Array Facility, Center for Integrative Genomics, University of Lausanne, CH-1015 Lausanne, Switzerland. Sylvain.Pradervand@unil.ch
authors.forename: Sylvain
authors.initials: S
authors.lastname: Pradervand
authors.forename: Johann
authors.initials: J
authors.lastname: Weber
authors.forename: Jérôme
authors.initials: J
authors.lastname: Thomas
authors.forename: Manuel
authors.initials: M
authors.lastname: Bueno
authors.forename: Pratyaksha
authors.initials: P
authors.lastname: Wirapati
authors.forename: Karine
authors.initials: K
authors.lastname: Lefort
authors.forename: G Paolo
authors.initials: GP
authors.lastname: Dotto
authors.forename: Keith
authors.initials: K
authors.lastname: Harshman
mesh_terms: D020869:Gene Expression Profiling / Q000379:methods*; D006801:Humans; D016014:Linear Models; D035683:MicroRNAs; D046228:Microarray Analysis / Q000379:methods*; D012680:Sensitivity and Specificity
chemical_list: D035683:MicroRNAs