Repurifying DNA - (Apr/11/2012 )

Hello!

First of all, I'm new to molecular biology, so many apologies if I'm full of questions. Second, this is something of a stupid question, so apologies again!

I just finished Maxiprepping, and I accidentally eluted in TE instead of EB. I need to send these samples for sequencing, but I know the EDTA in TE prevents the sequencing reactions from working correctly. How do I go about repurifying the DNA in water or EB so that I may send it out for sequencing?

We do have a PCR purification kit (the Wizard SV kit from Promega), can I use that?

Thank you!

You can ethanol precipitate the DNA or you could use the PCR purification kit (check the size cut-off), which is essentially an ethanol/isopropanol precipitation with a glass filter to retain the DNA.

Incidentally, I would have done the sequencing on a much smaller prep (e.g. mini) rather than scaling all the way up to a much more expensive maxiprep, before checking.

In my experience, TE has little or no effect on sequencing reactions, despite warnings to avoid it. It should not, since the EDTA is only 1 mM, and the amount of DNA in the reaction is typically 10-20% of reaction volume.

All right, thank you both!