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pup VS adult mice tail DNA - (Aug/27/2008 )

Hi
I got about 1.2ug/ul of genomic DNA while used adult mouse tail but with the same kit and all conditions for the mouse pup (3 weeks of age) i get just 50-100ng/ul.

why is it so.Is there any special precautions or additional steps while exttracting from a mouse pup.
I used Qiagen DNeasy tissue kit.

-bactin2-

QUOTE (bactin2 @ Aug 28 2008, 07:52 AM)
Hi
I got about 1.2ug/ul of genomic DNA while used adult mouse tail but with the same kit and all conditions for the mouse pup (3 weeks of age) i get just 50-100ng/ul.

why is it so.Is there any special precautions or additional steps while exttracting from a mouse pup.
I used Qiagen DNeasy tissue kit.


You should get buckets loads more DNA from a pup than from an older mouse. Perhaps you are overloading your columns?
Try again with less pup tail smile.gif
Clare

-Clare-

QUOTE (Clare @ Aug 28 2008, 08:44 PM)
QUOTE (bactin2 @ Aug 28 2008, 07:52 AM)
Hi
I got about 1.2ug/ul of genomic DNA while used adult mouse tail but with the same kit and all conditions for the mouse pup (3 weeks of age) i get just 50-100ng/ul.

why is it so.Is there any special precautions or additional steps while exttracting from a mouse pup.
I used Qiagen DNeasy tissue kit.


You should get buckets loads more DNA from a pup than from an older mouse. Perhaps you are overloading your columns?
Try again with less pup tail smile.gif
Clare


Oops,really?
Since i was not getting much DNA,i tried to put in more tail tissue.

-bactin2-

I would use less than 0.5cm from a 3 week old pup. I just extract the DNA by digesting with proteinase K and then doing a phenol/chloroform extraction. I would resuspend in 500ul water and only need to use 1ul in PCRs.

Good luck biggrin.gif

PS: Happy Friday!!!

-Clare-

You only need a tiny bit of the tail (I used less than 1mm) for plenty of DNA for PCR. Don't cut off larger bits of the tail, it hurts the mice and prevents them using it to climb in their cages. Too much tail will also result in having a lot of protein (including DNases) in the extraction, which can be very hard to remove if you are not careful.

Try not using a kit to extract, methods like phenol/chloroform extraction are much better and give really good yield if done properly.

-bob1-