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Automated RNA Amplification for Arrays

uantity is the labor-intensive column purification of cDNA and aRNA. Therefore the automation of mRNA preparation hinges on a robust and dependable method for purification. Whereas the standard column purification method requires extensive manual intervention, Ambion's MEGAclear magnetic bead purification technology is ideal for automation. The incorporation of a magnetic particle purification step after the cDNA synthesis step and then again after the in vitro transcription (IVT) step, allows aRNA sample preparation to be reformatted into a standard, automation-friendly 96 well plate.

GeneXP has validated and scaled up the automation of Ambion's Amino Allyl MessageAmp aRNA Amplification and 96 well MEGAclear purification system. The development of precise liquid-handling protocols allows maximization of purity and yields by careful control of RNA binding to and elution from magnetic beads. Consequently, it is now possible to amplify and purify diverse samples on multiple 96 well plates in the same two-day period that previously produced only 1020 aRNA preparations. This constitutes a greater than 10-fold improvement in throughput with high purity and high amplification yields, and with weekly throughput rising to 1,200+ samples per week on standard robots.

Figure 1 demonstrates the yield (Panel A) and purity (Panel B) of eight replicates of 2 g Promyelocytic Leukemia, Human Liver, and Universal Human Reference total RNA samples that were amplified and purified on two plates, for a total of 48 different samples. Samples were prepared as described by the Amino Allyl MessageAmp aRNA Amplification Kit protocol modified for robotic dispensation of re
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