Lambda DNA extraction protocol

Robin Beech robin_beech at maclan.mcgill.ca
Wed Dec 4 08:33:42 EST 1996


In article <32A4B910 at gate1.pbi.nrc.ca>, slee at pbi.nrc.ca ("Lee, Sun") wrote:

>Would anyone like to introduce me a really simple way for this?  Most of 
>published methods that were armed with 'easy' ,  'simple' ....   were 
>actually not like that.    Looking for in -hand method that really works.
>

Lambda miniprep

1) Grow an overnight liquid lysate
2) Spin down 1ml of lysate to remove bacterial debris
3) Transfer 0.8ml of lysate to a new eppendorf
4) Add DNAse to 1ug per ml and incubate at RT for 15 min
5) Add 0.2ml of Tris (50mM pH 8.0) EDTA (10mM) SDS (5%) shake gently and
incubate 70 deg for 15 min
6) Cool on ice and add 0.15ml 8M potassium acetate
7) Spin down the protein precipitate
8) Phenol-chloroform extract the supernatent
9) Add 2.5 vol of ethanol or 1 vol of isopropanol to precipitate the DNA

This can also be scaled up to process 8ml of lysate
Hope this helps - good luck

Robin N. Beech
Institute of Parasitology
McGill University



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