[Histonet] DNA extraction from wax sections

[Patsy Ruegg]

Isolation of DNA from paraffin-embedded tissue
DNeasy Tissue Kit Handbook  05/2002

Notes:
* lysis time will vary from sample to sample depending on the type of tissue
processed.
* Yields will depend both on the size and the age of the sample processed.
Reduced yields compared to fresh or frozen tissues are to be expected.
Therefore, eluting the DNA in 50-100 ul buffer AE is recommended.
* 1.  Place a small piece (not more than 25 mg) of paraffin-embedded tissue
in a 2 ml microcentrifuge tube.
* 2.  Add 1200 ul xylene.  Vortex vigorously.
* 3.  Centrifuge at full speed for 5 min. at RT.
* 4.  Remove supernatant by pipetting.  Do not remove any of the pellet.
* 5.  Add 1200 ul absolute ethanol to the pellet to remove residual xylene
and mix gently by vortexing.
* 6.  Centrifuge at full speed for 5 min at room temperature.
* 7.  Carefully remove the ethanol by pipetting.  Do not remove any of the
pellet.
* 8.  Repeat steps 5-7 once.
* 9.  Incubate the open microcentrifuge tube at 37 dC for 10-15 min until
the ethanol has evaporated.
* 10. Resuspend the tissue pellet in 180 ul Buffer ATL and continue with the
"DNeasy Protocol for Tissues".
* From this point the tissue can be treated as if it were fresh.

-----Original Message-----
From: histonet-admin <@t> lists.utsouthwestern.edu
[mailto:histonet-admin <@t> lists.utsouthwestern.edu]On Behalf Of Michele
Ellender
Sent: Thursday, November 06, 2003 8:55 AM
To: 'histonet <@t> lists.utsouthwestern.edu'
Subject: [Histonet] DNA extraction from wax sections

Dear all,
Does anyone out there have a protocol to extract DNA from sections of
formalin fixed wax embedded mouse tissues?
I've been asked to microdissect GI tumours from wax sections and extract the
DNA for use in PCR techniques.
Hope someone can help.
Many thanks,
Michele

Dr Michele Ellender
Radiation Effects Department,
NRPB, Chilton, Didcot,
Oxon. OX11 0RQ.  UK

michele.ellender <@t> nrpb.org



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