T4 PNK

peter via methods%40net.bio.net (by peter.ianakiev from gmail.com)
Tue Jun 24 10:41:18 EST 2008


On Jun 24, 2:02 am, d... from no.email.thankstospam.net (DK) wrote:
> In article <8bd38140-d8fa-49b5-a790-af29f4484... from m73g2000hsh.googlegroups.com>, peter <peter.ianak... from gmail.com> wrote:
>
>
>
> >On Jun 23, 9:58 pm, d... from no.email.thankstospam.net (DK) wrote:
> >> In article <mailman.543.1214259080.3533.meth... from net.bio.net>,
> > hroyc... from nmsu.edu wrote:
> >> >Good question!  I think long time ago ... in the last century ... Maniatis
> >> >et al.'s Lab Manual described the specificity of these enzymes.  I have
> >> >not really used dNTP for phosphoryltion, so I don't know how efficient it
> >> >would be.  Structurally the two are similar enough.
>
> >> I've never heard of any kinase being able to use dATP with any
> >> reasonable efficiency. I'd think that evolution worked out
> >> a way for things being  pretty good at distinguishing oxy vs
> >> and desoxy ribose. Much like DNAse vs RNAse.
>
> >> DK
>
> >I thought so too, but it seems that DNA end repair by T4DNA pol +
> >T4PNK works fine just with dNTP (not supplemented with rATP). Strange
> >Peter
>
> How exactly do you define "works fine"? And what kind of DNA
> is being end repaired? If it's genomic sheared DNA I would have
> thought just T4 poly would suffice, no need for PNK.

Lambda sonicated DNA jugged by shift after self ligation. Also  PCR
product produced by Taq pol and blunted, then self ligated.
-Peter


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