Preparation of genomic DNA from frozen tissue
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1st day:
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200...500 ng (about 1/4 cm3) tissue sample stored at - 80°C
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add 1 ml lysis buffer (20 mM TRIS pH 8; 1mM EDTA; 100mM NaCl; 0.5% SDS)
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add 20µl (= 0.3 mg) Protease K
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shake ON at 55°C
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2nd day:
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2 x extraction with equal volume PhOH/CHCl3
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2 x extraction with equal volume CHCl3
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ppt. with equal volume Isopropanol/NaOAc at RT
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30 min centrifugation
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wash with 1 ml 70% EtOH
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air dry
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resolve in 200 µl 1 mM TRIS pH 8 and shake for 2 hours at 4°C
to resolve the DNA
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2 µl on 1% TAE agarose gel for quality control and quantification
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