Self-cleaving sequences in Cloud Lab, possible?

General Science Discussion
1

We heard that switches will come back soon in EteRNA, but it’s still a few rounds away (so I expect that it will be a few more than that). In order to keep the crowd entertained (*evil grin*), I was wondering about experimenting with active sequences, like self-cleaving ones.

So, assuming that a sequence length N has a good potential to self-cleave at position K, what I would expect is that already at the end of the transcription phase, or after the heating/cooling step with proper refolding, in any case some time before the chemical mapping, we are left with the segment [K+1,N] which still should have the barcode and the long tail. If I’m not mistaken, this should still allow for reverse transcription and the rest of the mapping procedure.

But what happens after?

  • would these shortened sequences be screened and rejected as noise on ground of their length?
  • if not, what would appear on the SHAPE data for positions [1,K]? absolute reactivity? absolute non-reactivity? other?
2

Yup, should be possible. Note that the data for 1,K will end up being the SHAPE reactivity for non-cleaved RNAs only; and data for [K+1,N] will be the SHAPE reactivity for all RNAs. The data at K itself will end up being the fraction cleaved during our treatment, to a first approximation, I think. It might look kind of funny in the game.

So let’s try it! And then a fun exercise would be to include the self-cleaving sequence but design secondary structure that disallows the self-cleavage. [This is kind of similar to a puzzle that we had earlier this year with theophylline-induced switching of a hammerhead ribozyme – we’ll be reviving this soon, but I bet your puzzles would inspire rapid progress…]

3

Thanks Rhiju :slight_smile:
Well, I guess I have no choice now, I’ll prepare a couple projects on the topic :slight_smile:

4

And there goes my night (4am here)…
http://eterna.cmu.edu/web/lab/3509312/