Normalization: Introduction

As it is difficult to accurately determine the amount of RNA used for each array in the time course experiment, normalization of the data obtained from different array experiments for a direct comparison is critical for adapting DNA microarray to the quantitative analysis of mRNA turnover. However, the commonly used normalization method has been developed based on the assumption that there is no net change in overall gene expression (constant mRNA levels)(1). Although this default method has been proved to be successful for most DNA microarray experiments monitoring the relative mRNA levels of specific genes in two different samples, it is not applicable to mRNA decay analysis. This is because that the overall mRNA level is expected to decrease globally over time in these experiments. To solve this problem, we developed a new normalization method by doping a pool of in vitro transcribed polyA tagged Bacillus subtilis mRNAs with known concentrations into both RNA samples and reference samples as an internal standard for quantitative normalization.

References:

1. M. B. Eisen, P. O. Brown, Methods Enzymol 303, 179-205 (1999).