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MessageAmp II: Use Less RNA for Array Analysis

from RNA sample to sample.

Figure 2. Size of cRNA Produced from Different RNA Inputs. Bioanalyzer electropherogram of amplified RNA (single round amplification) derived from 1.0 and 0.1 g input HeLa cell total RNA. The mRNA Smear Assay was run in order to calculate the average size distribution. The center red line is the midpoint of the curve at 1030 nt for the 1.0 g input trace. The 0.1 g input midpoint size was 980 nt.

Figure 3. cRNA Derived From Different Total RNA Inputs. Bioanalyzer electropherogram of amplified RNA derived from 1000, 100, 50, and 10 pg input HeLa cell total RNA (two rounds amplification). The mRNA Smear Assay was run in order to calculate the average size distribution. The center red line is the midpoint of the curve at 630 nt for the 10 pg input trace and varies from 450 to 630 nt for the input range shown.


It is often the case that total RNA yield from a set of tissue or cell samples varies considerably. Some samples may yield 1 g of total RNA such that only one round of amplification is needed, whereas others may only yield 50 ng, thus requiring two rounds of amplification. For consistency within and between experiments, all samples in the study should be amplified with either one or two rounds of amplification; i.e. the amplification parameters for the smallest amount of input RNA in the sample set should be chosen for the entire study. Figure 4 shows some empirical data that may be useful in deciding whether one or two rounds of amplification are needed for samples smaller than 100 ng. In general, one round of amplification will not yield sufficient amounts of aRNA i
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