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Is RNA Amplification Necessary for Microarrays?

amplified RNA" were ranked by P-values, and 24 genes (2.1% of the total 1,150) were selected from the entire range of P-values for validation by qRT-PCR. Similar regulation patterns between array and qRT-PCR analysis were observed in 16 of the 24 genes tested.

(C) Six from the category of "genes unique to unamplified RNA" were validated by qRT-PCR. Of these, three genes showed a similar expression ratio by qRT-PCR analysis.

Thus amplification improved the sensitivity of the array analysis, enabling the identification of biologically regulated genes that might have been missed if unamplified RNA had been used.

Figure 1. Venn Diagrams Showing the Differential Distribution of Genes Expressed With and Without TNF-α Treatment. Of the total 13,284 genes tested, 1,296 were identified from amplified RNA vs. 146 genes that are commonly represented in both samples.

Table 1. Microarray Results. This study involved the comparison of multiple replicate arrays for each condition tested: (+TNF-α and TNF-α) with use of four unamplified and five amplified RNA samples. The gene expression ratios between microarray and quantitative RT-PCR data is presented in this table.


The Feldman Experiments
Feldman et al. evaluated the fidelity of microarrays probed with amplified vs. total RNA by comparing microarray data with real-time qRT-PCR. They compared gene expression in two murine tumor cell lines (B16F10 and
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