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ankalita 11-02-2012 08:23 PM

DNA digestion, ligation
 
Hi everyone,
I am facing problem at transformation step. After ligation I am not able to get the colonies. I change buffer, media and enzyme. Start getting colonies but those are not positive colonies. I have done digestion with Hind111 and XHO1. at 37 for 2hr then purified by qiagen gel extraction and ligate overnight at 4 oC.
My all grown positive colonies are showing smearing. What could be the reason. How I can troubleshoot this problem.


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