Systematic comparison of the fidelity of aRNA, mRNA and T-RNA on gene expression profiling using cDNA microarray

In cDNA microarray technology, there are three main reverse transcription based RNA labeling methods, using total RNA (T-RNA), mRNA, and amplified antisense RNA (aRNA), respectively. However, despite the common use of the three types of RNAs, limited data are available regarding their differences and concordances. In this report, we compared the three methods through two sets of self-comparison experiments using the same RNA sample in all cases. Within each method, duplicate hybridizations are highly reproducible with low biases, which are randomly produced. When combining different RNAs within a single array, correlation coefficients between the two channels are rather low, while the discrepancies are persistent. Furthermore, the fidelity of aRNA and mRNA microarrays in the expression profile study shows no significant difference with standard T-RNA based labeling methods. These results suggest that some RNA abundance are selectively changed during aRNA amplification/mRNA purification processes, but it will not affect the gene expression ratio of the two samples if the same type RNA are used. Therefore all three types of RNAs can be used in expression profiling analysis as long as the test and reference samples are generated by identical method within single study.