Team:University College London/Protocols/Electrophoresis

From 2012.igem.org

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'''Preparing the Gel'''
'''Preparing the Gel'''
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1. Within a conical flask, add 3ml 50X TAE, 1.5g Agarose, and 150ml RO water
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Step 1: Within a conical flask, add 3ml 50X TAE, 1.5g Agarose, and 150ml RO water
-
2. Heat for 1 min in microwave. Swirl. Heat again for 30s. If solution is clear stop. Else repeat.
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Step 2: Heat for 1 min in microwave. Swirl. Heat again for 30s. If solution is clear stop. Else repeat.
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3.Cool solution under running cold water.  
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Step 3: Cool solution under running cold water.  
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4. Add 20ul ethidium bromide (normal concentration of EB solution is 500ug/ul)
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Step 4: Add 20ul ethidium bromide (normal concentration of EB solution is 500ug/ul)
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5. Pour into a sealed casting tray in a slow steady stream, ensuring there are no bubbles
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Step 5: Pour into a sealed casting tray in a slow steady stream, ensuring there are no bubbles
'''Running a gel'''
'''Running a gel'''
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1. Add 1 part loading buffer to five parts of loading sample
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Step 6: Add 1 part loading buffer to five parts of loading sample
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2. Position the gel in the tank and add TAE buffer, enough to cover the gel by several mm
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Step 7: Position the gel in the tank and add TAE buffer, enough to cover the gel by several mm
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3. Add 5ul of DNA ladder to lane 1
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Step 8: Add 5ul of DNA ladder to lane 1
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4. Add samples to the remaining wells
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Step 9: Add samples to the remaining wells
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5. Run at 100 volts for 1hour and 15 minutes
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Step 10: Run at 100 volts for 1hour and 15 minutes
'''Imaging the Gel'''
'''Imaging the Gel'''
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1. Place gel in GelDoc 2000 chamber
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Step 11: Place gel in GelDoc 2000 chamber
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2. Turn GelDoc 2000 chamber on 
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Step 12: Turn GelDoc 2000 chamber on 
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3. From computer: Quantity One > Scanner > Gel_Doc_Xr>Manuqal Acquire
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Step 13: From computer: Quantity One > Scanner > Gel_Doc_Xr>Manuqal Acquire
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4. Alter the exposure/settings to give a clear image.
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Step 14: Alter the exposure/settings to give a clear image.
TAE - Tris-acetate-EDTA
TAE - Tris-acetate-EDTA

Revision as of 08:45, 1 August 2012

Electrophoresis

Preparing the Gel

Step 1: Within a conical flask, add 3ml 50X TAE, 1.5g Agarose, and 150ml RO water

Step 2: Heat for 1 min in microwave. Swirl. Heat again for 30s. If solution is clear stop. Else repeat.

Step 3: Cool solution under running cold water.

Step 4: Add 20ul ethidium bromide (normal concentration of EB solution is 500ug/ul)

Step 5: Pour into a sealed casting tray in a slow steady stream, ensuring there are no bubbles

Running a gel

Step 6: Add 1 part loading buffer to five parts of loading sample

Step 7: Position the gel in the tank and add TAE buffer, enough to cover the gel by several mm

Step 8: Add 5ul of DNA ladder to lane 1

Step 9: Add samples to the remaining wells

Step 10: Run at 100 volts for 1hour and 15 minutes

Imaging the Gel

Step 11: Place gel in GelDoc 2000 chamber

Step 12: Turn GelDoc 2000 chamber on 

Step 13: From computer: Quantity One > Scanner > Gel_Doc_Xr>Manuqal Acquire

Step 14: Alter the exposure/settings to give a clear image.

TAE - Tris-acetate-EDTA EDTA - ethylenediamine tetraacetic acid

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