storing RdRp at -20C

D.K. dk at no.email.thankstospam.net
Sun Mar 14 12:37:20 EST 2004


On 14 Mar 2004 05:57:06 -0800, alion85 at hotmail.com (redeamer) wrote:

>Hi guys!
>
>in a few days i am up to control my RNA dependent RNA polymerase for
>its activity, i have dialyzed it against Tris-based buffer, 50%
>glycerol, 1mM EDTA,
>1mM DTT, 0,01% TritonX-100, 50mM NaCl(may be ive skipped
>something)...and i put a 5mM MgCl2.. i know that EDTA will chelate
>some 2mM of my MgCl2, 

1 mM. Each EDTA binds one Me2+. 

>and i know that in general PCR we should add
>MgCl2 (that is discrete)to DNA polymerases..
>
>My question is whether anybody of You knows is it critical for
>maintaining an active RdRp to store it with a Mg2++ or otherwise? Or
>its very personal for each particular RdRp, as one could suppose?

Not sure of this articular case but for most Mg-dependent enzymes
there is no significant difference. Cases where Mg binding also 
plays a structural role in stability are not that frequent. 
That said, if there is no Me2+-dependent protease activity, there 
is nothing wrong with keeping Mg around.

DK




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