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PCR of PCR product not working

PCR of PCR product not working - Molecular Biology Techniques

PCR of PCR product not working - Molecular Biology Forum. Includes forums for common molecular biology techniques.


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Old 04-03-2013, 02:50 PM
Pipette Filler
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Question PCR of PCR product not working



Hi
I am not getting enough concentration of my amplified gene in PCR reaction for cloning purpose. I am trying to increase concentration of my amplified gene by doing rePCR of my PCR product by using PCR product as a template, but this doesn't work. I would like to know is PCR of PCR product would work or not? also how to increase the concentration of gene of interest by PCR as I got thin band of my gene of interest which may or may not be sharp and also got other bands. So I use gel extraction kit method to purify my gene of interest but there is too much loss of product and some times I do not get visible product and enough concentration. Also PCR of such purified product again gives multiple bands which I am not expecting so it is difficult to me to increase concentration of my gene of interest for cloning.

please suggest

thanks

Ajit
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Old 05-05-2013, 03:27 PM
SMG SMG is offline
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Default Re: PCR of PCR product not working

if you are using boiling method please use commerical kits. Sometimes it is helpful to get DNA. But if you already used it, you should check primers, i had same problem, i had to add one more primer to see band profile.
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