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MessageAmp II: Use Less RNA for Array Analysis

ng more than 1 g of HeLa S3 RNA does not significantly increase aRNA yield. Also, kidney RNA, which generally produces less aRNA than other tissues, does not produce proportional increases in aRNA yield with increasing amounts of input RNA up to above 1 g. Figure 1 also provides a useful benchmark for understanding the relationship between the amount of aRNA produced and the source and quantity of input RNA.

Figure 1. MessageAmp II aRNA Amplification Yields. Yields of aRNA (g) amplified using the MessageAmp II aRNA Amplification Kit from six different RNA sources using total RNA inputs of 503000 ng. Yields shown are the average of duplicate reactions. The IVT incubation time was 4 hours for all samples. Purified aRNA concentrations were measured using a NanoDrop spectrophotometer.

*Universal Human Reference RNA (Stratagene)


The IVT reaction time also has an impact on aRNA yield from MessageAmp II reactions. The yield of aRNA increases with increasing RNA input and IVT time. As a rule, most amplification reactions containing between 1001000 ng input total RNA, incubated overnight (14 hr IVT), will produce sufficient aRNA for any microarray platform (at least 10 g). While a minimum of 4 hours incubation time is recommended, yields obtained from 23 hour incubations are often sufficient, and >500 ng input RNA in a 24 hr incubation will typically produce enough aRNA for microarray analysis.


One or Two Rounds of Amplification?
Based on experimental requirements and
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