genomic DNA extraction from formalin fixed tissues (not embedded) for genotyping - (Jul/13/2012 )
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
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.
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!
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.ac.uk/68731/1/Palero_SciMar_2010.pdf
2) This one looks more promising, use normal chemicals (DNA extraction from formalin fixed Dolphin tissues):
http://www.zoo.ba.gov.br/biblioteca/veterinaria/dna_extraction.pdf
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 : )