Gel analysis of ligation product
(by sjvinay from yahoo.com)
Tue Nov 11 05:28:19 EST 2008
I have a question about gel analysis of ligation product. I would like to insert a 1.7 Kb fragement into a 3.3 Kb vector. I have done the restrictions. I would like to do a gel analysis before transforming to save me time with mini preps. I have done around 10 ligations so far. On the gel, I see that some times I get blanks(i.e no vector, no insert and no ligated products). Some times I get ladders of various combinations of vectors and inserts. These ladders do not transform and I do not get any colonies. Now, my question is if the blanks are the optimal ligation products ? I read in his forum that since ligated products have varying winding numbers, they are not to be seen on a gel. Can anyone confirm this ?
I know that nicked circles migrate slowly on a gel and I was expecting a slowly moving band, which I was planning on purifying and transforming. But so far I have not been able to get this.
Any help would be appreciated.
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