IGEM:UBC/2009/Notebook/UBC iGEM 2010/2010/06/22

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DspB Track

Marianne and Vicki

Gel Verification

  • Verifying PCR products from last day (PCR-ing genomic preps)
  • See Gel Verification Protocol in "iGEM Training Workshop Molecular Biology 2010"

Procedure:

  1. Prepare one 0.8% agarose gel
    1. Weigh out 0.8g of agarose and transfer to a 250mL flask
    2. Add 100mLs of 1x TBE buffer and swirl gently to disperse the agarose
    3. Microwave the mixture on high power until it boils and the agarose is completely dissolved.
    4. Allow the solution to cool to 60ºC by incubating in a 60ºC waterbath for about 10 minutes
    5. Add the appropriate (5uL per 50mL) amount of stain; in this case: add 10uL of GelStar
  2. Pour the cooled agarose into the plate with the appropriate number of combs
  3. Wait approximately 20 minutes
  4. Transfer 2uL of each PCR sample into new microfuge tubes
  5. Add 8uL of dH2O to each PCR sample
  6. Use quick and dirty way of loading gel:
Pipet 2uL of DNA loading buffer for each of the samples onto parafilm
Pipet 10uL of sample, then pipet the DNA loading buffer (2uL) for a total of 12uL, then load onto gel

Note: Used 100bp DNA ladder, a 1 in 20 dilution
Machine Conditions: 100V, 45 minutes, 1xTBE buffer Gel arrangement:

Table 1. Arrangement of gel
ABCDEFGHIJ100bp DNA ladderKLMNOPQRS


Note: May have added more water in sample C -MP
Results:
File:100622 DspB 1.png

Lab Stuff

  • Autoclaved 10uL x2 boxes of pipette tips, microcentrifuge tubes 1.7mL x2 bins