PCR and Proteinase K ?

theodorn at medlib.georgetown.edu theodorn at medlib.georgetown.edu
Fri Aug 28 17:42:09 EST 1998


In article <MGALgAALwm51EwFu at demon.co.uk>,
  "Dr. Duncan Clark" <duncan at genesys.demon.co.uk> wrote:
> In article <Pine.GSO.3.96.980827115611.13926B-100000 at cmgm.Stanford.EDU>,
> Markus Schneemann <schneema at cmgm.Stanford.EDU> writes
> >If yes - do you know a trustworthy, pure, DNAse-free Proteinase K for our
> >purposes ?
>
> Could you not just incubate the proteinase K on it's own for an hour at
> 55C to get it clean of DNAse? In the old days one used to sit Pronase at
> 37C for two hours prior to using it for the first time.
>
> Duncan
<sig snipped>

That Pronase comment really takes me back, Duncan.  Thanks for the trip down
memory lane. It looks like we've both dated ourselves.

In regards to Proteinase K; it really is clean (at least the stuff from
Boehringer is [no affiliation]).  I haven't been "pre-digesting" it, and I
use it routinely for cytoplasmic RNA extraction, and haven't had a whiff of
RNA degradation (even after once forgetting a tube in the water bath and
letting my RNA digest in Proteinase K overnight).  I have also gotten into
the habit of removing CIAP with Proteinase K after phosphatasing DNA , so it
appears to be DNAse-free, too.

Nick Theodorakis

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