removing nucleic acids from extracts
knut at iti.org
Wed Jan 31 10:02:14 EST 1996
In article <310CD910.4A12 at quickmail.ucsf.edu> trey simmons <trey_simmons.gicd at quickmail.ucsf.edu> writes:
>of nucleic acid is left, which is interfering with subsequent steps.
>know there must be a straightforward way to remove nucleic acids
it is common to precipitate the dna with either streptomycin sulfate (1% final)
or polyethylenimine (0.5% final). the streptomycin sulfate can be made up
as a 5% solution, and neutralized (not always necessary). then add 1/5 vol
dropwise while stirring in the cold. stir for ~1 hour, then spin at 10k rpm
in sorval ss34. if you have a lot of plasmid, it might not all come down.
if the solution is cloudy after centrifugation, not all the dna precipitated.
one can add more, and spin again. the resulting supernatant should be clear,
with a color from yellow to light brown, depending on the ammount of protein.
any milky cloudyness is dna that has not been precipitated. pretty much
the same protocol can be used for polyethylenimine, except that one would
add 1/20th volume of a 10% solution. again, if the resulting supernatant
is not clear, add more and spin again.
--knut knut at rosa.mit.edu
More information about the Proteins