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I am working on an (expensive) synthetic construct, which happens to have many "repetitive" sequences within it that are integral to its function. Primarily, the two sequences that are worrying me are:

  • A long polyA (about 15 nucleotides).
  • A consecutive sequence of two nucleotides: GGGUUUGGUUGU, etc.

I've heard from many some of the transcription horrors they've been through because of similar sequences with similar nucleotides. However, some advise that transcribing it shouldn't be a problem.

How predictably can RNA polymerase transcribe these sorts of sequences in E.coli?

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1 Answer 1

I don't think repetitive sequences are intrinsically problematic to an E coli. Bacteria also deal with things like polyadenylation. Basically, you have nothing to lose from trying -- E coli is one of the more robust expression platforms, and it's difficult to predict whether the sequence will fold into some tertiary structure (which can easily impede translation.)

So I think the predictability should be set at the baseline for E coli RNA polymerase error rate, which is about $10^{-5}$.

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