RNA in plasmid preparation interfere in ligation - RNA contamination and ligation (Apr/16/2002 )
I have prepared plasmid (PET3a) in large scale and found lot of RNA contamination. But i am scared to RNAse treatment for i may loose some amount of plasmid DNA. Can i go ahead with RE digestion and ligation without RNA treatment. Because anyway i will be running my REdigest in electrophoresis and following extraction before ligation.
Thanking in advance for you suggestions.
If your RNase is clean you won't need to worry about loosing any of your plasmid. Are REs inhibited by RNA? THat's a good question. To avoid having to answer it just perform an RNase A treatment. No worries..
You will almost certainly be safe treating your DNA with
RNaseA. Add an excess of the enzyme, then treat @ 37C for 1 hour. Phenol chloroform extract, then ethanol precipitate.
Works every time for DNA from Midi-preps etc etc