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Ligation: two basic questions


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#1 Spright

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Posted 21 February 2013 - 03:02 AM

1. If the number of colonies after a ligation matches the number on the negative control (no insert), is there still a chance that some of the colonies are actually good?

2. I have a mixture of two PCR products that I'd like to ligate in tandem, one connected to each other, into a vector. However, one of them is capable of ligating into the vector alone. Is this ligation even worth trying? Will there be 100% preference for the small single insert, or is there rather a good chance that I obtain what I wish?

#2 GNANA

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Posted 21 February 2013 - 04:13 AM

To your first question, theoritically NO , practically its bit unlikely, but if you are lucky you may !!!!!!

2.ya nothing wrong in trying though the chances of getting what you wish is less, may be you might have to screen, screen and screen lot of colonies to get. Alternatively you can try a sequential ligation, like ligating the pcrs first and then adding it on to the ligation mix with vector alone, in this way you can increase the chances.

good luck....
I would prefer being perfectionist rather than a passionist in Research.

I always had an alternate hypothesis....




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