Preparation of 5'monophosphate mononucleotides from double stranded DNA

• 1st day:
• 2...5 µg dsDNA of high quality  in solution which must not contain EDTA
• add 10 µl 10 x Exonuclease III buffer (Boehringer) and adjust to 98 µl
• 5 times 2 sec sonication (15%)
• add 1 µl ExoIII (Boehringer) and 1 µl Nuclease P1 (1g/l)
• incubate ON at 37°C
 
• 2nd day:
• heat to 95°C for 5 min to denature remaining double-stranded fragments
• add 1 µl 3M NaOAc pH 5.2 and 1 µl Nuclease P1
• incubate at 37°C for 1 h and 72°C for 1 hour

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