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Bacteriophage genome extraction

Bacteriophage genome circular permutation DNA sequencing genome extraction

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#1 Hamoba

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Posted 17 January 2013 - 03:57 PM

Hi all,

We have recently a bacteriophage which appears to be a Siphoviridae. It has got a small genome of about 20 Kbp. I isolated the genome with standard phenol/ chloroform extraction and ethanol precipitation but after extraction and when I want to digest, the genome seems to be degraded and smear on the gel. I checked all the reagents used in the experiment but they all seem to be fine and work with other + controls. Even if I incubate the extracted DNA in water at 37 C it smears on the gel and if I add a restriction enzyme buffer without the RE and incubate at 37 the DNA gets completely degraded and nothing is visible on the gel. I was just wondering if anybody knows why this is happening and maybe have had similar experience and how to resolve the issue.

Many thanks.

#2 phage434

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Posted 17 January 2013 - 06:32 PM

Did you redissolve the DNA after precipitation in TE? Resuspension in water can lead to rapid degradation.





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