DNA extraction from polyacrylamide gels

Errol E.S.Kwan at massey.ac.nz
Sat Aug 26 22:36:42 EST 2000


It may be possible that the amount you extracted was not enough to be
detected by ethidium bromide but would suffice for PCR.  So far I have
not had any problem extracting DNA from a polyacrylamide gel.

Errol

On Fri, 25 Aug 2000 10:02:22 -0500, Lise Larsen
<larsenli at missouri.edu> wrote:

>I've tried  the Qiaex II Gel Extraction Kit (Qiagen) with good results.
>Thanks for the response I've received.
>
>To Vince Mulholland: My gels are stained with ethidium bromide. I tried to
>concentrate the DNA extracted by freeze/thawing and run it on an agarose gel
>without prior reamplification, just to see if I got any DNA out of my
>extraction. I didn't so I concluded that the problem was the extraction
>rather than the following PCR amplification.
>
>Lise
>
>Vince Mulholland wrote:
>
>> Lise,
>>
>> Rather than your extraction method, it could be to do with the staining
>> method you use. Do you silver stain your DGGE gels? If you silver stain,
>> do you use a stop solution which contains acetic acid? The acetic acid,
>> after silver staining, seems to render DNA refractory to amplification
>> (presumably due to nick of the strands).
>>
>> Vince
>>
>> Vincent Mulholland,
>> Senior Plant Pathologist,
>> Diagnostics & Molecular Biology Section,
>> Scottish Agricultural Science Agency,
>> East Craigs,
>> Edinburgh EH12 8NJ, U.K.
>>
>> URL:    http://www.sasa.gov.uk/
>> E-Mail: Vince.Mulholland at sasa.gov.uk
>> Tel:+44(131)2448845 Fax:+44(131)2448912
>>
>> ___________________________________________________
>> How do you extract DNA from a polyacrylamide gel?
>>
>> I run DGGE gels and need to excise the bands for PCR reamplification and
>> sequencing. I've tried cut out bands and freeze/thaw them ×3 in water
>> but didn't get any product after the following PCR. Please, let me know
>> if you know of a good method to recover the DNA.
>>
>> Thank you
>> Lise Larsen
>>
>> ---
>>
>> ---
>






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