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Genomic DNA Preparation from RNAlater Preserved Tissues


The following protocol is suitable for 100 mg tissue samples. Mouse kidney, lung, and spleen genomic DNA have been successfully extracted with approximate yields of 70 g DNA per 100 mg tissue.

Required Materials

Razor blade or scissors 55C incubator
Nuclease-free 2.0 ml microfuge tubes Nuclease-free water
1 M Tris, pH 8.0 Buffer saturated phenol
0.5 M EDTA Chloroform
10% SDS 3 M Sodium acetate, pH 5.2
Proteinase K 95% Ethanol


Protocol

  1. Prepare digestion buffer:
    60 mM Tris, pH 8.0
    100 mM EDTA
    0.5% SDS

  2. Remove tissue from RNAlater and mince well. Cutting the tissue into very small pieces is absolutely essential to achieving adequate digestion. Place in a 2.0 ml eppendorf tube with 1.5 ml digestion buffer.

  3. Add Proteinase K to a final concentration of 500 g/ml. Mix by inversion. Incubate the sample at 55C for approximately 4 hours
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