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Slack / Private Message Drop, p.999 [SLACK_001213] · slack_pm:msg:10427

Date: 2021-09-21 08:14
Type: chat message · slack
recipient: Robert F. Garry, Edward C. Holmes, Andrew Rambaut
speaker: Kristian G. Andersen

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Again, it depends on cloning strategy - if the use restriction enzymes and there would be an easy way to insert anFCS with PCR utilizing REs already present in the sequence, then it could be a way to do it. Not the optimal way, for sure, but sometimes ease trumps perfection.

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2021-09-21 08:10 Andrew Rambaut open

Key question is - would anyone chose to put the FCS we see in SC2 in and put it out of frame.
2021-09-21 08:11 Kristian G. Andersen open

Would be a very strange way to do it, but not impossible - depending on their cloning strategy.
2021-09-21 08:12 Andrew Rambaut open

If you wanted to insert an FCS you would pick the AAs you wanted, pick the codons you wanted and then insert it.
2021-09-21 08:12 Andrew Rambaut open

The alternative is that it was inserted in frame and then mutated to make it look out of frame
2021-09-21 08:14 Kristian G. Andersen

Again, it depends on cloning strategy - if the use restriction enzymes and there would be an easy way to insert anFCS with PCR utilizing REs already present in the sequence, then it could be a way to do it. Not the optimal way, for sure, but sometimes ease trumps perfection.
2021-09-21 08:16 Andrew Rambaut open

Would you pick CGG for example? Would you consider what codon to use for an R or just pick one arbitrarily.
2021-09-21 08:21 Kristian G. Andersen open

CGG would be an unusual choice IMO - not a 'optimized' R and sorta middle-of-the-pack as it comes to usage in human genomes. Very rare, of course, in viruses because it's a CpG.
2021-09-21 08:24 Andrew Rambaut open

And you have 6 codons to chose from.
2021-09-21 08:25 Kristian G. Andersen open

Correct. Key here too is that if you were to 'optimize' the FCS sequence (as is commonly done for these types ofexperiments) you would _not_ end up with the FCS sequence we see, including the CGG codons. But again, itdoesn't really tell us much.

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