4 replies to this topic

[Roo]

Does anyone have a good protocol for isolating genomic DNA from formalin fixed tissues, preferably without using a kit?  I need to repeat genotyping of some mice that are now stored in formalin.  I understand that long term storage in formalin can greatly decrease my success rate, but I don't have fresh or frozen tissue to use so I have no choice.  

I found protocols using kits, salt precipitation, and various digest buffers, but most are for paraffin embedded samples.  I don't have to do a xylene step to get rid of the paraffin, so I'm thinking I could try picking a protocol and starting after the xylene step.   However, I don't know if I should perform wash steps and/or the drying steps that are found in so many protocols.  I may just try something since I don't have anything to lose, but I was hoping somebody might have a tried and true method that usually works for their lab.

Any information or suggestions would be greatly appreciated!

-Roo

Edited by Roo, 13 July 2012 - 07:39 AM.

[Adrian K]

There is one article:
Analytical Biochemistry 395 (2009) 265–267   doi:10.1016/j.ab.2009.08.016

This protocol using mineral oil, not xylene. See if this paper helps.

[Roo]

Thanks, but this article describes the use of mineral oil as a deparaffinization agent.  It was used in conjunction with commercial DNA extraction kits.  My samples aren't embedded in paraffin.  They're just mouse carcasses sitting in formalin.    For the time being, I can't justify buying a DNA extraction kit for just a few samples when I already have most things needed for extraction.  I was simply looking for a protocol that someone has personally used.  Thanks for trying though!  I will probably end up doing a bit of trial and error.  If it works, I'll post it!

[Adrian K]

I'm sorry as I overlooked your query... my bad. However, based on Google search, here are 2 papers which might help you.

1) This protocol uses Tetramethylsilane (TMS)-Chelex (not sure if you have this chemical in your lab)
http://eprints.soton...SciMar_2010.pdf

2) This one looks more promising, use normal chemicals (DNA extraction from formalin fixed Dolphin tissues):
http://www.zoo.ba.go..._extraction.pdf

[Roo]

Thanks Adrian!  The second article is perfect!!!  It's similar to others that I found, but better.  I probably won't do it this week, but I'll let you know how it turns out.  Thanks again!

-Roo : )