Team:UNIPV-Pavia/Methods Materials/Electrophoresis

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(Difference between revisions)
(→Electrophoresis)
(→Electrophoresis)
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'''Ethidium bromide'''<hr color='white' width='70%'>
'''Ethidium bromide'''<hr color='white' width='70%'>
'''Loading buffer 10x Blue Juice, Invitrogen'''<hr color='white' width='70%'>
'''Loading buffer 10x Blue Juice, Invitrogen'''<hr color='white' width='70%'>
-
'''TBE (Tris/Borate/EDTA buffer) 5x (final volume 1 liter)'''<hr color='white' width='70%'>
+
'''TBE (Tris/Borate/EDTA buffer) 5x (final volume 1 liter)'''
-
*'''54 gr Tris'''
+
**'''54 gr Tris'''
**'''27.5 gr Borate'''
**'''27.5 gr Borate'''
'''20 ml EDTA 0.5 M (pH 8)'''<hr color='white' width='70%'>
'''20 ml EDTA 0.5 M (pH 8)'''<hr color='white' width='70%'>

Revision as of 16:32, 22 September 2009

EthanolPVanimation.gif



Protocols


Electrophoresis

(estimated time: 2 hours)

  • Prepare agarose gel in 1x TBE buffer
  • Add ethidium bromide (using gloves and face mask for your safety):
    • 1 µl in the small size agarose gel (70 ml)
    • 2 µl in the middle size agarose gel (150 ml)
    • 4 µl in the big size agarose gel (250 ml)
  • Cast the gel, insert the well-forming comb and let it polymerize
  • Add the loading buffer to each sample
  • Load the samples and 8 µl of marker
  • Set to 70-100 volts and electrophorese for the required amount of time
  • Use UV-light to look at the bands
  • Take a picture of the gel, if needed (not when bands have to be cut!!!)










DNA samples
Ethidium bromide
Loading buffer 10x Blue Juice, Invitrogen

TBE (Tris/Borate/EDTA buffer) 5x (final volume 1 liter)

    • 54 gr Tris
    • 27.5 gr Borate
20 ml EDTA 0.5 M (pH 8)
Marker (1 kb DNA Ladder, Promega)
Face mask and gloves
Electrophoresis apparatus

Transilluminator





  • NOTE: when not specified, the marker has the following pattern:
Pv ladder.jpg
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