IGEM:UNAM/2009/Notebook/IGEM project/2011/05/31

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Extract the plasmid PBBMRCS-5
Abstract Later I did a agarose gel of the plasmid PBBMRCS-5, with the purpose of seeing if the extraction was cleaned of chromosome.
 * Today I extracted plasmid PBBMRCS-5 with the method of alcaline lysis.

Method to extract plasmid DNA


 * 1. Prepare two eppendorf tubes of Escherichia Coli DH5-α.
 * 2. Centrifuge the tubes to 13 rpm during three minutes and decant the supernatant.
 * 3. Add TE 10, shake the tubes with the vortex and centrifuge.
 * 4. Remove the supernatant with a syringe.
 * 5. Put 100 μl of the solution 1 to he cells and shake it.
 * 6. Put 200 μl of the solution 2 and mix for inversion and centrifuge for ten minutes to 13 mil rpm.
 * 8. Put 200 μl of the solution 3 and shake for inversion.
 * 9. Pass the supernatant to a new eppendorf tube.
 * 10.Put 1 ml of ethanol to 100% in the new eppendorf tube and shake with the vortex.
 * 11.Centrifuge to 13 mil rpm for ten minutes and remove the liquid with the syringe.
 * 12.Put 1 ml of ethanol to 70% and shake with the vortex.
 * 13.Detach the cell pill of the tube and centrifuge.
 * 14.Dry the the cell pill with the speed vac.
 * 15. Put 20 μl of TE-RNASA.

Gel of the plasmid PBBMRCS-5


 * Gel of 1 gr. of agarose and 100 ml of TAE.


 * Line 1st -> Green ladder 5 μl.
 * Line 2nd -> PBBRMCS-5 5 μl + 2 μl of dye. (1)
 * Line 3rd -> PBBRMCS-5 5 μl + 2 μl of dye. (2)
 * Line 4th -> PBBRMCS-5 5 μl + 2 μl of dye. (3)
 * Line 5th -> PBBRMCS-5 5 μl + 2 μl of dye. (4)
 * Line 6th -> Green ladder 5 μl.