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[mythy]

Hi,

I recently purified plasmid DNA using the Qiagen plasmid purificaiton kits and also using a protocol for alkali lysis (Birnboim and Doly).

When I ran the Qiagen plasmid DNA on a gel as well as the Birnboim and Doly plasmid, I found a sharp band for Birnboim and Doly whereas the Qiagen kit plasmid showed a smear.
The Qiagen smear was localised around the position of plasmid DNA, and at lower UV exposure two sharp bands were visible in the smear.

Spectrophotometric analysis showed that there were comparable amounts of DNA in the two samples, and both samples were loaded in the same way, so I don't see how Qiagen could be overloaded while Birnboim was not.
To test this, I ran the gels again with a quarter dilution of the samples and also loaded less total volume onto the gel. This resulted in the same smearing for Qiagen, while Birnboim samples were not visible.

This seems to suggest that there is a difference in the conformations of the plasmid DNA and DNA fragments between the two samples.

Any suggestions?

Thankyou