Hello all,
I am wondering if anyone knows of any algorithms or work that has been done on rational design of mRNA for increased stability/expression (in eukaryotic cells). I know that AU rich regions in the 3`UTR of mRNA can target them for destruction, thus resulting in short half-lives, but I haven't seen a lot of work published on optimizing RNA for increased half-life (at least in eukaryotes).
Thanks,
J
Thanks for the answers. Andrei, I have used such services before by other companies (GenScript). Do you know if there has been a comparison between Life Technologies algorithms and GenScripts?
Also, a part of the reason for asking the questions isn't just to get higher expression levels, but to know if RNA can be rationally designed to alter its half life. For instance, generate a series of mRNA transcripts that all have different half lives.
Chemical modifications to RNA (e.g. methylation) can stabilize in vivo and in vitro transcribed mRNA. There is a nice recent review in Nature Reviws Genetics http://www.nature.com/nrg/journal/v15/n5/full/nrg3724.html
Export of spliced mRNAs (incorporated in expression vectors) from the nucleus can be a problem and export can be enhanced by adding WPRE (woodchuck hepatitis post-transcriptional regulatory element) to the construct.
If you want a set of similar mRNA transcripts with different stability, you might try to render them substrates for nonsense-mediated mRNA decay (NMD) for which decay efficiency correlates with the position of the stop codon in the mRNA.
The introduction of nonsense mutations in the ORF (which will result in truncated peptides of course) at different positions or increasing the length of the 3'-UTR (not altering the encoded proteins) results in increasing destabilisation of the mRNA (up to some point). Please check https://www.researchgate.net/publication/5404055
I hope this might help you.
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