Isolated plasmid

==Isolating Plasmid from Cells (Miniprep)==

* Pellet overnight culture by centrifuging 13,000g for 10 minutes. 
* Pipette out supernatent and discard. 
* Add 250μl resuspension buffer R3 and mix by pipetting up and down. 
* Transfer to capped tubes. 
* Add 250μl lysis buffer L7 and mix by inverting tube 5 times. 
* Incubate for 5 minutes at room temperature. 
* Add 350μl precipitation buffer. Invert until mixture is homogenous. 
* Centrifuge 13,000g for 10 minutes. Place spin column into recovery tube. 
* Load supernatent into spin column and discard capped tube. 
* Centrifuge 13,000g for 1 minute. Discard supernatent. 
* Add 700μl wash buffer W9 (with ethanol). 
* Centrifuge 13,000g for 1 minute. Discard supernatent. 
* Centrifuge 13,000g for 1 minute to remove all liquid. Discard any remaining supernatent and recovery tube. 
* Place spin column in new recovery tube. 
* Add 100μl TE buffer or MilliQ H2O. 
* Incubate for 1 minute at room temperature. 
* Centrifuge 13,000g for 2 minutes. The supernatent in the recovery tube should contain isolated plasmid. 
* Discard spin column and store plasmid at -20°C.

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