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removing foreign DNA


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#1 lyok

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Posted 31 July 2013 - 01:36 AM

Dear all,

 

I have been working on a DNA extraction and one of the steps involves to remove the unwanted foreign DNA by destryoing it and destroying bacterial cells in the sample with bleach. 

The bleach itself does not damage my cells so its easy to use, however during the bleach step, the protocol states to keep the sample with the bleach on ice. Why is this? Doesnt the bleach work at room temperature?

Or is it just to prevent that the chlorine evaporates and thus making the bleach worthless ?

 

Or is it also to prevent my cells to be damaged? Which seems weird..

 

 

any other ideas?

I was thinking about preventing DNAses to become active.. but they have already been removed by a previous step in which they have been destroyed already.



#2 bob1

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Posted 31 July 2013 - 02:22 AM

Bleach does work at room temp and would probably kill your bacteria if left for long enough.  However, I suspect the temperature will be to ensure that your cells are not too damaged, but most (all?) of the contaminating ones are disrupted by the incubation.  Doing this at room temp would be difficult as the incubation period would probably be very short and a small error likely to damage your cells as well.



#3 lyok

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Posted 31 July 2013 - 02:53 AM

Bleach does work at room temp and would probably kill your bacteria if left for long enough.  However, I suspect the temperature will be to ensure that your cells are not too damaged, but most (all?) of the contaminating ones are disrupted by the incubation.  Doing this at room temp would be difficult as the incubation period would probably be very short and a small error likely to damage your cells as well.

So it works better at room temperature! Ok good to know. 

I was thinking the chlorine would evaporate too fast at room temperature, since they always state to store bleach at cold temperatures. 



#4 bob1

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Posted 31 July 2013 - 12:32 PM

The usual bleach solution is sodium hypochlorite, which slowly decomposes in water to release Cl, this process is quite slow and the bleach should be stable for quite a while at RT.  Household bleach solutions (usu 3-8% hypochlorite) are stored at room temp for months (at least mine lasts that long) with no loss of activity that I can see.



#5 lyok

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Posted 25 August 2013 - 01:53 AM

The usual bleach solution is sodium hypochlorite, which slowly decomposes in water to release Cl, this process is quite slow and the bleach should be stable for quite a while at RT.  Household bleach solutions (usu 3-8% hypochlorite) are stored at room temp for months (at least mine lasts that long) with no loss of activity that I can see.

Ok thanks.






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