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DNA Precipitation


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#1 sanu

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Posted 26 May 2010 - 01:00 PM

Hi,
After following the protocol for DNA precipitation according to the following protocol, if I do not see any pellet does that mean that I do not have any DNA?

dd 0.1 volume 3 M sodium acetate
add 2.5 volumes 100 % Ethanol
vortex
precipitate at:
-20C overnight (+++)
-80C 1 h (++)
dry ice 15min (+)
spin 20 minutes at 12000 rpm 4C
carefully pour out / aspirate supernatant (do not lose DNA-pellet)

I used dry ice.I did the precipitation and out of six tubes, only one had a visible pellet. Does that mean all others have no DNA??

Please help.

Thanks,
Sanu

Edited by sanu, 26 May 2010 - 01:25 PM.


#2 moerae

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Posted 26 May 2010 - 02:11 PM

Hi,

Do you expect to have a lot of DNA? If not, then the lack of pellet doesn't mean there's no DNA. The DNA pellets I've worked with are usually very glassy and sometimes very hard to spot. I'd just keep going and resuspend and do a Nanodrop or run a gel of whatever you've got and see if there's anything. No point throwing it out when you've done most of the hard work already.

Hope that helps.

#3 sanu

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Posted 26 May 2010 - 06:24 PM

Hi,

Do you expect to have a lot of DNA? If not, then the lack of pellet doesn't mean there's no DNA. The DNA pellets I've worked with are usually very glassy and sometimes very hard to spot. I'd just keep going and resuspend and do a Nanodrop or run a gel of whatever you've got and see if there's anything. No point throwing it out when you've done most of the hard work already.

Hope that helps.


I agree. But in one of the tubes, I saw a huge white pellet and the others no pellet at all. So I was wondering if there is no DNA. I will proceed with the ligation and seee if that works. Thanks for your input. It is really of help.




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