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Checking DNase activity


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#1 floyd78

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Posted 25 September 2012 - 12:25 PM

We have had a minor catastrophe where a freezer completely defrosted over the weekend so everything was brought up to room temperature for an unknown amount of time.

I need to use Ambion Turbo DNA-free on some total RNA preps I have stored elsewhere but I want to assess whether the defrosted DNase and accessory buffers are still useable as it was a new unopened kit.

Does anyone have a protocol for assessing DNase 1 activity, preferably using electrophoretic methods as access to the Bioanalyzer is currently limited.

Many thanks

#2 Curtis

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Posted 27 September 2012 - 08:30 AM

I just know that sometimes DNase works without its original buffer. I have tried.I remember using just 1 ul of DNase on a big pellet of genomic DNA at 50-60C for 10 min and it digested it. So maybe you don't even need to check its activity.

#3 tgb

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Posted 28 September 2012 - 12:45 AM

We have had a minor catastrophe where a freezer completely defrosted over the weekend so everything was brought up to room temperature for an unknown amount of time.

I need to use Ambion Turbo DNA-free on some total RNA preps I have stored elsewhere but I want to assess whether the defrosted DNase and accessory buffers are still useable as it was a new unopened kit.

Does anyone have a protocol for assessing DNase 1 activity, preferably using electrophoretic methods as access to the Bioanalyzer is currently limited.

Many thanks

Hi floyd78,
There is a simple solution for you. We used it when we had similar problem.
You need PCR product. It is not important their size or anything.
Incubate PCR product with your DNAase kit in proper condition. You can try to add your enzyme in different concentration.
When the incubation time end, load your samle on the gel.
If you don't see any band on the gel that's mean your kit is still working.

#4 floyd78

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Posted 28 September 2012 - 01:09 AM

Many thanks for everyones help. I've managed to find some genomic DNA that shows no signs of degradation so I'll have a look today using the suggested protocols.




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