enzyme digestion of a PCR product

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roxanalibe
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enzyme digestion of a PCR product

Post by roxanalibe » Thu Jul 15, 2010 9:07 am

Hello,


I need to cut with EcoR I and XhoI a PCR product in which I introduced the restriction sites for these enzyme and I am wondering if I should cut with both of them in the same time and purify from the gel, and then perform ligation, or I should cut with one at a time and then purify after each step. What do you think?

Thank you for your input!!!

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JackBean
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Post by JackBean » Thu Jul 15, 2010 9:12 am

go to http://www.lmgtfy.com/?q=double+digest and pick your provider's page to look, whether these two enzymes can be used together and in which buffer

Based on NEB's page they can be used together and in almost all buffers, but check your provider ;)
http://www.biolib.cz/en/main/

Cis or trans? That's what matters.

roxanalibe
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Post by roxanalibe » Thu Jul 15, 2010 10:02 am

thank you, I already used the double-digestion NEB site, but I was thinking that in the case of just a PCR product is different from a plasmid, for which I usually dont have any problem to use double-digestion.

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JackBean
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Post by JackBean » Thu Jul 15, 2010 12:25 pm

that shouldn't be problem. But you should rather do the PCR, gel purification and then digestion.
http://www.biolib.cz/en/main/

Cis or trans? That's what matters.

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