I suppose you could try the ligase cycling reaction with synthesized oligos then do a PCR off of it. Since Taq ligase can't add nucleotides, literal positioning could accomplish directionality. (have one overlap 20 with the sgRNA, one overlap 25, etc, and design the sgRNA to also have this)
-- I've also looked into goldengate assembly, it can do this pretty darn well.
-Koeng
On Tuesday, August 11, 2015 at 4:42:54 AM UTC-7, Mega [Andreas Stuermer] wrote:
On Tuesday, August 11, 2015 at 4:42:54 AM UTC-7, Mega [Andreas Stuermer] wrote:
Yeah, but the problem is that multiple repeats are difficult to synthesize for synthesis companies, and some won't even try. Or make it much more expensive.
On Monday, August 10, 2015 at 11:37:16 PM UTC+2, Michael Flynn wrote:Hi all,I recently wrote my undergraduate thesis on RNA secondary structure prediction algorithms. In the process I've become intimately familiar with the algorithms and code bases behind Unafold and RNAstructure, which I have been told comprise 90% of the market share of RNA structure prediction software. My thesis is located here, if you are interested.However, I was a physics and computer science student, and I have minimal (no) training in biology. This meant that most of the time while I was writing my thesis I was in great want of background knowledge and motivation. I was often asked why my research was important, but I could never give a strong answer. When I asked my advisor for motivations behind our work, he did not answer to my satisfaction. So I thought, what better way to learn than DIYbio?So are there ways I could help this community? What would be the best path for me to get started, in a way that my background would be the most relevant?Mike
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