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DNA precipictation at -80 went thick

DNA precipitation

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#1 Marigold77

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Posted 20 December 2017 - 09:23 AM

Hi all,

 

I've just done normal DNA precipitation in a 50ml falcon (DNA in 10ml H20, 0.1 volume 3M NaAcOH, 2.5 volumes 100% Ethanol) and placed it at -80 for 3h. When I came to take it out it was all super thick. Has anyone seen anything like this before? This never happened to me at -20.

My DNA pellet yield after centrifugation was tiny so I left it at -20 now in hope that whatever did not precipitate at -80 due to the fact that the reaction went weirdly thick, will precipotate at -20 as usual. I'd appreciate any input.

 

 

 



#2 merlav

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Posted 20 December 2017 - 11:55 AM

When I do DNA precipitation I use only the absolute alcohol (ethanol or isopropanol) and the NaAcOH, I don't add water.  I let it on -80C for a few hours or -20c O/N.  Then centrifuge, decant, 5mL of 70% EtOh (prepared with nuclease free water) flick or tap the tube, centrifuge, decant, place over a kimwipe,  then left it inside the biological cabinet or PCR working station for 10 min to dry all the alcohol. Resuspend with TE1x


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#3 Trof

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Posted 10 January 2018 - 07:34 AM

Is it supposed to be kept in -80 C?

This table shows freezing point of 70% ethanol alone at -48 C. I'd guess it is freezing so doesn't precipitate well then.

https://www.engineer...ater-d_989.html


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