Purifying Recombinant plasmid DNA

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xbritoe
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Purifying Recombinant plasmid DNA

Post by xbritoe » Mon Jan 16, 2012 8:23 am

Hi, so I have been transforming recombinant plasmid DNA into E.coli bacteria to clone the DNA. The plasmid consist of an ampicillin resistant gene. After I grow the bacteria, I would purify the plasmid DNA out of the bacterial cell using a maxi prep kit. So I was wondering if the final product of my purification consist of plasmid DNA (including the AMP resistant gene) or does it only consist of the DNA fragment that I initially placed into the plasmid.


I have been using my cloned DNA to transfect a cell line, so I was just wondering if I have been transfecting it with only my target gene or if I have been transfecting the whole plasmid including the AMP-R gene.

Thank you

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JackBean
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Post by JackBean » Mon Jan 16, 2012 9:10 am

the kit extracts any intact DNA molecule of particular size, thus you will obtain whole plasmid
http://www.biolib.cz/en/main/

Cis or trans? That's what matters.

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JackBean
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Re: Purifying Recombinant plasmid DNA

Post by JackBean » Mon Jan 16, 2012 9:12 am

xbritoe wrote:I have been using my cloned DNA to transfect a cell line, so I was just wondering if I have been transfecting it with only my target gene or if I have been transfecting the whole plasmid including the AMP-R gene.

Shouldn't you be the one who knows? Depends on what cell line and what was the protocol, you could either use whole plasmid or only part of it. But the transformation was probably based on recombination, thus you should have been transfecting only your gene plus something extra.
http://www.biolib.cz/en/main/

Cis or trans? That's what matters.

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