Minipreps: alkaline lysis based plasmid purification

Описание: Aka alkaline lysis, the “miniprep” is a technique in which we separate and purify the plasmid DNA we put into bacteria (the DNA we want) from all the stuff that was already in the bacteria (the DNA we don’t want, proteins, sugars, etc.). You can’t just break the cells open (lyse them) & pull out all the DNA because the bacteria has its own DNA which you aren’t interested in. But some cool chemistry comes to the rescue!

full text & figures on blog: http://bit.ly/minipreps

The basic workflow (once you've amplified the plasmid by growing the bacteria in some liquid media (food)) is:
resuspension: spin cultures to pellet cells, pour off the media and put them in better conditions
alkaline lysis: EDTA & SDS help break the cells open, NaOH disrupts base-pairing to unzip the DNA
neutralization: potassium acetate lowers the pH to allow re-zipping (which plasmid's small enough to do okay) but the genomic DNA strands can't find their match before they're dragged out with the potassium-SDS-coated insoluble gunk
purification: pellet out all that gunk, move the liquid (supernatant) to a spin purification column where the DNA will bind, wash the salts off while the DNA's bound, then get the DNA to unbind and elute (come out) in water or elution buffer

here are links to DIY versions:

https://openwetware.org/wiki/Qiagen_B... &

http://malooflab.phytonetworks.org/wi...