need ~100 bp fragments from Saliva DNA

Jose de las Heras via methods%40net.bio.net (by josenet from tiscali.co.uk)
Sun Sep 16 08:51:27 EST 2007


"Ozan Aygun" <metugenetics from yahoo.com> wrote in message 
news:mailman.962.1189902662.11350.methods from net.bio.net...
> Hi,
>
> I suggest you to try partial digestion with
> Micrococcal nuclease or Benzonase.
>
> Treatment with Benzonase decreases viscosity very
> quickly when incubated >20 C and has a broad range
> buffer compatibility. You can check the size of the
> fragments in a time course titration experiment with
> benzonase incubation & concentration and optimize your
> desired DNA size.
>
> Hope this may be any help, good luck!
>
> Ozan

I'd be careful with micrococcal nuclease (and use small amounts [1]). Its 
suitability depends on the intended use for the DNA. This enzyme will not 
just nick the DNA, but it'll chew away the ends. Since you're starting from 
purified DNA (no proteins bound) you may assume that the enzyme will create 
nicks and digest DNA totally at random locations, so this could be okay... 
but I'd test it first if you need to be sure that you don't have sequence 
biases.

[1] I've used it to help fragment DNA in crosslinked chromatin samples. In 
order to ensure reproducibility I ended up performing the digestions at 
0-4oC, because I found that digestion at 37oC was too quick for my purposes.

Jose 




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