IGEM:UNAM/2009/Notebook/IGEM project/2011/05/30

From OpenWetWare

Jump to: navigation, search
iGEM Project name 1 Main project page
Previous entry      Next entry

Gel of PBBMRCS-5 plasmid

Abstract

Today I did a gel of the DNA plasmid PBBMRCS-5, the gel showed that the plasmid was cleaned of chromosome.

  • 1. I did the agar to 1%.
  • 2. I put the gel in running buffer and fill the wells of the gel.
  • 3. Volume of substances in each well of the gel:
  • 1st lane -> 5 μl of green ladder.
  • 2nd lane -> 5 μl of plasmidic DNA PBBRMCS5 + 2 μl of loading dye.
  • 3rd lane -> 5 μl of plasmidic DNA PBBRMCS5 + 2 μl of loading dye.
  • 4th lane -> 5 μl of plasmidic DNA PBBRMCS5 + 2 μl of loading dye.
  • 5th lane -> 5 μl of green ladder.
  • 4. I put the gel in the BIO-RAD during 110 minutes to 90 Volts.
  • 5. I washed the gel with distilled water.
  • 6. I put 100 μl of etide bromure miligramo/mililitro with distilled water in a tray and put the gel. Wait 10 minutes.
  • 7. I flushed the gel with water and put in distilled water during 10 minutes.
  • 8. I saw the gel in the GBOX.

Note: PBBMRCS-5 was extracted using alcaline lysis and 15μL Plasmid DNA digested with XBA I and Hind III.

PBBRMCS-5 Plasmid Digestion

Reagent Quantity
H2O 2.5 μL
NEBuffer 2 10X 2.5 μL
XBA I 1.25 μL
Hind III 1.25 μL
BSA 2.5 μL
PBBRMCS-5 Plasmid DNA 15 μL
Total 25 μL

Leave all night at 37ºC in agitation.



Personal tools