Hello! I'm doing CTAB DNA extraction from some fungal groups, but sometimes I have some salt crystals residuals, and when I send PCR products to sequenciate, it doesn't work.
I don't know why they appear, because I have extracted DNA from the same individual several times, and one of those time appear salt residual.
How can I clean it when appears? Or I have to extract again DNA?
Thank you!
María José
Plasmid Clean-Up for removing salt from backbone of DNA:
Hello María José,
do you perform a PCR on the extracted DNA and send the PCR product? If yes, the salt contamination is more likely to arise from the PCR mix. I can recommend a PCR clean up step either by column or gel extraction.
If this is not the case I would recommend an additional 70% ethanol wash as I assume you extract with isopropanol? Salt and DNA precipitate, but not in ethanol.
Hi María José. I think I know what happened to you cause we had the same problem many times. The critical step is the isopropanol precipitation. If you add too much isopropanol, you get CTAB precipitation together with DNA. I you use the exact volume isoprop from your protocol, you have to pay attention to the volume of supernatant you previously transfered. we sometimes reduce this volume so we don't get solid materials from the interface, and due to this reduction, we perform the precipitation with excess of CTAB. So the solution is, you either add CTAB solution to the pipeted supernatant, or yoy reduce the volume of isoprop in the next step. both work. For each ml of supernatant you want to use 0.7 to 0.75 ml cold isoprop. i'm sure this is going to work. don't hesitate to contact me in case of doubts!
bst
Francisco
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