Nucleic Acids Research

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Nucleic Acids Research, 2002, Vol. 30, No. 4 e15
© 2002 Oxford University Press

Normalization for cDNA microarray data: a robust composite method addressing single and multiple slide systematic variation

Yee Hwa Yang¹, Sandrine Dudoit², Percy Luu³, David M. Lin³, Vivian Peng^3,4, John Ngai^3,4,5 and Terence P. Speed^1,6,*

¹Department of Statistics, ²Division of Biostatistics, ³Department of Molecular and Cell Biology, ⁴Functional Genomics Laboratory and ⁵Neurogenomics Center, Helen Wills Neuroscience Institute, University of California, Berkeley, CA 94720-3860, USA and ⁶Division of Genetics and Bioinformatics, The Walter and Eliza Hall Institute, Melbourne, Australia

There are many sources of systematic variation in cDNA microarray experiments which affect the measured gene expression levels (e.g. differences in labeling efficiency between the two fluorescent dyes). The term normalization refers to the process of removing such variation. A constant adjustment is often used to force the distribution of the intensity log ratios to have a median of zero for each slide. However, such global normalization approaches are not adequate in situations where dye biases can depend on spot overall intensity and/or spatial location within the array. This article proposes normalization methods that are based on robust local regression and account for intensity and spatial dependence in dye biases for different types of cDNA microarray experiments. The selection of appropriate controls for normalization is discussed and a novel set of controls (microarray sample pool, MSP) is introduced to aid in intensity-dependent normalization. Lastly, to allow for comparisons of expression levels across slides, a robust method based on maximum likelihood estimation is proposed to adjust for scale differences among slides.

^* To whom correspondence should be addressed at: Department of Statistics, University of California, 367 Evans Hall, #3860 Berkeley, CA 94720-3860, USA. Tel: +1 510 642 2781; Fax: +1 510 642 7892; Email: terry{at}stat.berkeley.eduThe authors wish it to be known that, in their opinion, the first three authors should be regarded as joint First Authors

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				M. G. Wade, A. Kawata, A. Williams, and C. Yauk Methoxyacetic Acid-Induced Spermatocyte Death Is Associated with Histone Hyperacetylation in Rats Biol Reprod, May 1, 2008; 78(5): 822 - 831. [Abstract] [Full Text] [PDF]

				R. Bhati, C. Patterson, C. A. Livasy, C. Fan, D. Ketelsen, Z. Hu, E. Reynolds, C. Tanner, D. T. Moore, F. Gabrielli, et al. Molecular Characterization of Human Breast Tumor Vascular Cells Am. J. Pathol., May 1, 2008; 172(5): 1381 - 1390. [Abstract] [Full Text] [PDF]

				R. A. Chavez Montes, P. Ranocha, Y. Martinez, Z. Minic, L. Jouanin, M. Marquis, L. Saulnier, L. M. Fulton, C. S. Cobbett, F. Bitton, et al. Cell Wall Modifications in Arabidopsis Plants with Altered {alpha}-L-Arabinofuranosidase Activity Plant Physiology, May 1, 2008; 147(1): 63 - 77. [Abstract] [Full Text] [PDF]

				T. del Castillo, E. Duque, and J. L. Ramos A Set of Activators and Repressors Control Peripheral Glucose Pathways in Pseudomonas putida To Yield a Common Central Intermediate J. Bacteriol., April 1, 2008; 190(7): 2331 - 2339. [Abstract] [Full Text] [PDF]

				K. Heurlier, M. J. Thomson, N. Aziz, and J. W. B. Moir The Nitric Oxide (NO)-Sensing Repressor NsrR of Neisseria meningitidis Has a Compact Regulon of Genes Involved in NO Synthesis and Detoxification J. Bacteriol., April 1, 2008; 190(7): 2488 - 2495. [Abstract] [Full Text] [PDF]

				A. Hausmann, B. Samans, R. Lill, and U. Muhlenhoff Cellular and Mitochondrial Remodeling upon Defects in Iron-Sulfur Protein Biogenesis J. Biol. Chem., March 28, 2008; 283(13): 8318 - 8330. [Abstract] [Full Text] [PDF]

				B. Guo, Y. Wang, F. Shi, Y.-W. Barton, P. Plummer, D. L. Reynolds, D. Nettleton, T. Grinnage-Pulley, J. Lin, and Q. Zhang CmeR Functions as a Pleiotropic Regulator and Is Required for Optimal Colonization of Campylobacter jejuni In Vivo J. Bacteriol., March 15, 2008; 190(6): 1879 - 1890. [Abstract] [Full Text] [PDF]

				A. T. Beek, B. J. F. Keijser, A. Boorsma, A. Zakrzewska, R. Orij, G. J. Smits, and S. Brul Transcriptome Analysis of Sorbic Acid-Stressed Bacillus subtilis Reveals a Nutrient Limitation Response and Indicates Plasma Membrane Remodeling J. Bacteriol., March 1, 2008; 190(5): 1751 - 1761. [Abstract] [Full Text] [PDF]

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