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In vitro Transcription for RNA Synthesis

In vitro synthesis of single-stranded RNA molecules is a widely used laboratory procedure that is critical to RNA research, as well as to RNA biopharmaceuticals. This technique is versatile in that it allows the researcher to tailor synthesis and introduce modifications to produce a transcript. Downstream applications include biochemical and molecular characterization of RNA for RNA-protein interactions and structural analyses, generation of RNA aptamers, synthesis of functional mRNAs for expression, and generation of small RNAs for alteration of gene expression (e.g., guide RNAs, RNAi). Furthermore, the use of in vitro synthesized RNA has been instrumental in the development of RNA vaccines and CRISPR/Cas9 genome editing tools, generation of pluripotent stem cells, as well as development of RNA amplification-based diagnostics.

High-yield robust IVT reactions require optimization of each reaction component. NEB offers six in vitro RNA synthesis kits, all of which have been optimized to generate reproducible yields of quality RNA. In addition, individual components can also be purchased for IVT and mRNA capping.

  1. Rio, D. C., et al. RNA: A Laboratory Manual. Cold Spring Harbor: Cold Spring Harbor Laboratory Press, 2011, 205-220.
  2. Cooper, Geoffery M. The Cell: A Molecular Approach. 4th ed. Washington D.C.: ASM Press, 2007. 262-299.
  3. Jani B, Fuchs R. J Vis Exp. 2012 Mar 26 (61). PMID: 22473375

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FAQs for In vitro Transcription for RNA Synthesis
Protocols for In vitro Transcription for RNA Synthesis
    Publications related to In vitro Transcription for RNA Synthesis
    • Wu, M.Z., Asahara, H., Tzertzinis, G., Roy, B. (2020) Synthesis of low immunogenicity RNA with high-temperature in vitro transcription RNA; PubMedID: 31900329
    • Eyler, D.E., Franco, M.K., Batool, Z., Wu, M.Z., Dubuke, M.L., Dobosz-Bartoszek, M., Jones, J.D., Polikanov, Y.S., Roy, B., Koutmou, K.S (2019) Pseudouridinylatio of mRNA coding sequences alters translation Proc Natl Acad Sci U S A; 116(46), 23068-23074.. PubMedID: 31672910, DOI: 10.1073/pnas.1821754116
    • Potapov, V., Fu, X., Dai, N., Correa, I.R., Jr., Tanner, N.A., Ong, J.L (2018) Base modificatons affecting RNA polymerase and reverse transcriptase fidelity Nucleic Acids Res; 46(11): 5753-5763, PubMedID: 29750267
Recommended HiScribe RNA Synthesis Kits by Application
RNA Polymerase Selection Chart
NEB offers RNA polymerases that can be used for in vitro synthesis of RNA for a wide variety of downstream applications. Additional polymerases are offered for the generation of untemplated homoribopolymeric tails for reverse transcription or labeling.
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