- Alvaro E. Rodriguez M. 21:45, 21 February 2013 (EST):It would be nice if you added literature examples of Directed Evolution of Proteins for each approach as done in the Ancestral Sequence Reconstruction or take a similar approach.
- Kevin Baldridge 16:43, 25 February 2013 (EST):Gotta give a plug for the technique my group likes secM ribosome display
- Kevin Baldridge 16:44, 25 February 2013 (EST):it would be nice to have a one or two sentence summary of the other techniques you list in the advanced/high-throughput section
- Gabriel Wu 16:55, 25 February 2013 (EST): For those of you who do this sort of thing, have you any thoughts on the contexts in which Directed evolution works well and, more importantly, where it doesn't work well? Since most papers are about successes, do you know of any examples (personal communication/experience) where it doesn't work and speculation on why?
- Evan Weaver 17:00, 26 February 2013 (EST): I remember you said what the genetics method was for high throughput and screening on monday, but what was it again? The list is very vague.
- Evan Weaver 17:03, 26 February 2013 (EST): What does improved codon usage mean in the context of synthetic GFP?
- Neil R Gottel 16:45, 28 February 2013 (EST): Different organisms will differ in the amount of each tRNA that corresponds to each codon. Certain codons are rare in some species, while common in others. So, if you're putting a jellyfish gene into E. coli, then the codon usage is likely not optimized. Then production of that gene's product will be slower/lower (because it takes longer to produce a peptide if the ribosome is waiting around for a rare tRNA to come by). However, according to this OWW page on Codon usage optimization, and specifically this paper, the most important factor to consider is which tRNAs are charged (that is, get amino acids attached to them) when the cell is starving, and to favor using the corresponding codons when optimizing your gene. I haven't actually done this sort of optimization though, so hopefully someone else more experienced can chime in.