Help with significant loss of DNA after extraction from gel

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5 years ago

Stefan Fuhrmann

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Help with significant loss of DNA after extraction from gel

Hello everyone. I am in need of some help. After isolating DNA and amplifying it I ran it through an agarose gel to check it. When I tried to extract it from the gel the amount of DNA I got was too low to use. For instance, I started with around 500 ng/μl but managed to extract only concentrations as low as 10 ng/μl. What might be causing this significant loss? Also, I am wondering if purifying the pcr product with PEG mix is sufficient.

Experiment: DNA gel extraction / PCR product purification Product size < 15Kb

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**Dr. Natasha Pasricha** · Answer 1 · 2019-05-23T13:08:53+00:00

Firstly, you should try to elute the DNA in low volumes of 10-20 μl so that you increase the concentration of the DNA while maintaining the same yield. Additionally, you might want to use a PCR clean up kit since the PEG based method might result in loss of your PCR product. I am fairly confident you can find some good PCR clean up kits here on Labettor. Finally, you can try to re-amplify your PCR product from your gel and purify it again.