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HomePolymerase Chain Reaction ApplicationsIntegration of Sigma® TransPlex® WTA and the Complete WTA2 Kits with the NimbleGen™ Expression Microarray Workflow

Integration of Sigma® TransPlex® WTA and the Complete WTA2 Kits with the NimbleGen™ Expression Microarray Workflow

Amplification products generated by the TransPlex® WTA and Complete WTA2 kits are suitable for microarray target for expression analyses, and can be readily integrated into existing NimbleGen workflows.

  • If your total RNA sample quantity is less than 1 microgram, amplify the experimental and control (input) samples using the Transplex WTA Kit (WTA1) or the Complete WTA2 Kit (WTA2-50RXN) before labeling.
  • The WTA amplification product ranges from ~200 to 1000 base pairs in size, averaging ~300 base pairs, making fragmentation unnecessary. This size is sufficiently large for NimbleGen microarray target.
  • Purify samples using the GenElute™ PCR Cleanup kit (NA1020).

Preparation of WTA Amplification Product for Labeling

  1. Perform WTA amplification as described in the product bulletins for the TransPlex WTA or Complete WTA2 kits, found on the Sigma-Aldrich website.
  2. Purify the amplification product using the GenElute PCR Cleanup kit (NA1020) eluting with sterile RNase-/DNase-free water (W4502 or W1754).
    Note 1. Thirty microliters are the absolute minimum elution volume Elute with 50 ul for maximum yield.
    Note 2. The capacity of the GenElute PCR Cleanup filter cartridge is 10 μg, sufficient for the typical yield of a single TransPlex WTA amplification reaction.
  3. If concentration of the amplification product is required, use vacuum-centrifugation to avoid loss of amplified product.
Materials
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References

1.
Special acknowledgement and thanks to Simon Melov, PhD, Associate Professor and Director of the Genomics Core, and Krysta Felkey, BS, Research Scientist, the Buck Institute for Age Research, Novato, CA, for their assistance in the preparation of this procedure.