IGEM:Harvard/2009/Notebook/Harvard iGEM 2010/2010/07/14: Difference between revisions

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[[Image:strep+mirabrazz_STOP gel.jpg|| 350px]]
[[Image:strep+mirabrazz_STOP gel.jpg|| 350px]]
    # 1kb Plus Ladder
# 1kb Plus Ladder
    # Miraculin C Strep E/S
# Miraculin C Strep E/S
    # Miraculin N Strep E/S
# Miraculin N Strep E/S
    # Brazzein C Strep E/S
# Brazzein C Strep E/S
    # Brazzein N Strep E/S
# Brazzein N Strep E/S
    # 1kb Plus Ladder
# 1kb Plus Ladder


==Team Fence==
==Team Fence==

Revision as of 12:29, 14 July 2010

iGEM iGarden <html><img src="/images/9/94/Report.png" border="0" /></html> Main project page
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Team Flavor

Miniprep of STOP + V0120 part

   Nanodrop O/D's:
   STOP V0120 1-1: 
   STOP V0120 1-2: 
   STOP V0120 2-1: 
   STOP V0120 2-2:

DTL of StrepII+Mira/Brazz with STOP codon

  1. 1kb Plus Ladder
  2. Miraculin C Strep E/S
  3. Miraculin N Strep E/S
  4. Brazzein C Strep E/S
  5. Brazzein N Strep E/S
  6. 1kb Plus Ladder

Team Fence

PCR of the arabidopsis promoters with the newly extracted DNA and the older DNA. FAIL

Arp2 promoter PCR failed.

Exp2 promoter PCR failed.

Phenol Chloroform DNA Extraction

  • resuspend DNA in approximately 200μL molecular grade pure water
  • add 1 volume phenol chloroform
    • when pipetting the phenol chloroform be sure to pipette from closer to the middle level of the liquid as the top two centimeters do not contain what we want
  • centrifuge at top speed for 5 minutes
  • collect the aqueous top layer and transfer to a clean microcentrifuge tube
    • the remaining liquid is toxic and must be carefully disposed of in the "toxic waste" bottle in the fume hood
  • add 1 volume phenol chloroform
  • centrifuge at top speed for 5 minutes
  • collect the aqueous top layer and put into a fresh tube
  • add three volumes 100% ethanol and 1/10 volume sodium acetate
  • allow solution to incubate on ice for 15 minutes
  • centrifuge at top speed for 10 minutes
  • pipette off the aqueous solution leaving a pellet of DNA in the bottom of the tube
  • wash in 500μL 70% ethanol
  • mix tube by inverting
  • centrifuge again for 10 minutes at top speed
  • air dry pellet by leaving tube open on bench (not on ice) til dry
  • after pellet has dried resuspend DNA in desired volume of H2O or buffer EB
    • presence of DNA can be confirmed by running a small sample on a gel

THE END


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