High-Yield Synthesis of RNA Using T7 RNA Polymerase and Plasmid DNA or Oligonucleotide Templates
Adapted from RNA: A Laboratory Manual, by Donald C. Rio, Manuel Ares Jr, Gregory J. Hannon, and Timothy W. Nilsen. CSHL Press, Cold Spring Harbor, NY, USA, 2011.Abstract
When a large amount (milligram quantities) of RNA is desired (e.g., for making RNAs for RNA interference studies or for structural analyses), it is advantageous to use optimized transcription conditions. The method described here uses high millimolar concentrations of rNTPs, pyrophosphatase, and a specially formulated high-yield transcription buffer. These conditions prevent product (pyrophosphate) inhibition of RNA polymerase, which permits the accumulation of large quantities of RNA.
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