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How can I improve my Gel Extraction yields. We use the standard protocol from Qiagen, gel extraction, dissolve in QG buffer at 42C and purify via anion exchange columns. However, with 500 ng we ultimately get 100 ng for a 20% yield. I was curious what else has been tried for gel extraction.

We also add IPA and NaOAc to the dissolved mixture prior to the column purification.

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3 Answers 3

When I optimized gel extraction in my hands, two factors turned out to be critical in maximizing gel extraction yield:

The pH of the eluting solution. I used to elute the final product with DEPC-treated water initially, but the yield would vary a lot as the pH of the water changed. Using buffer EB is better, since it maintains a stable pH 8.

Most importantly, *incubate the eluting solution with the column as long a possible (and also elute twice). * I incubate for about 10 min, which is longer than what they recommend but it works much better.

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I also want to use this post to mention there are cheaper alternatives to the Qiagen kits, such as the ones here: mn-net.com, that work just as well and sometimes better. Using those can save you a lot of money. This is not an ad: I or my lab are not in any way affiliated with the company or website I'm linking to. I'm just sharing customer experience. –  Drosophila Mar 22 '13 at 15:01

Try dissolving at 50C. In the Qiagen gel extraction kit, it says to dissolve at 50C or until completely dissolved.

The size of the gel is important too. Make sure it is 1.5% or less, and the size of the gel needs to be less than 400 mg.

Also, pay attention to the first step. If the liquid is not the same color as the color of the first QG buffer then there is a pH problem.

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Elute from the column with Elution Buffer that has been preheated to 50-60 degrees. I stick mine in the incubator for the duration of the extraction so it is warm when I come to elute. It seems to help yield.

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