Team:Michigan/Quorum Sensing

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Revision as of 20:58, 21 July 2010


Michigan Header




Sunday Monday Tuesday Wednesday Thursday Friday Saturday
Week 1 - - - - - - -
Week 2 - - - 7/7/2010 7/8/2010 - -
Week 3 - - - - - - -
Week 4 7/17/2010 - 7/19/2010 - - - -

Quorum Sensing Team

Members include Alex Pyden, Marcus Lehr, Jennifer Hong, Eric Raynal, Katie Miskovich, and Audra Williams

7/7/10

Jennifer and Alex - in Lin Lab

Made CaCl2 stock solution: 0.1M - 50mL

  • molar mass = 111 g/mol
  • (0.1 mol/L)(.05 L)(111 g/mol) = 555mg
  • dissolved 555 mg CaCl2 in 50mL DIwater
  • vacuum filtered into 50mL tube

Made ampicillin stock solution: 

 -1 g ampicillin
 -5mL DIwater
 -ethanol
  • filtered by syringe into 15mL tube
  • put into ten 1mL alliquots in 1.5mL tubes
  • stored in -20°C in ERB lab

Inoculated E. coli DH5α from frozen stock into 12 mL LB broth

  • put in 30°C, 200 rpm shaking, overnight

~1.5 hrs

7/8/10

Alex, Jennifer and Eric - in ERB

Made LB agar plates w/ ampicillin 

 -10 g LB broth
 -7.5 g agar
 -500 mL DI water
  • autoclaved mixture at 121°C for 30 min (sterilize time), following Mike Nelson's protocol

Created protocols for Obtaining Deionized Water in the ERB and ERB Spectrophotometer

poured 20 LB-amp plates (large)

  • left to solidify in ERB 1230
  • 12 plates were used by Ann for biobrick transformations
  • 8 plates were stored in 4°C ERB

~3.5 hrs work

Ann, Marc, Audra and Katie - in Lin Lab

Performed biobrick transformations using heat shock

After second washing:

  • ODs for sample 1 and 2 were 0.378 and 0.358 respectively

4 hrs of work

7/17/10

Alex & Jennifer - in ERB

Yesterday, Marcus stored newly obtained strains in 4°C in ERB

  • W3110 w/ plasmids pTC6 & pET-GFP
    • AI-2 reporter (E. coli, amp and kan-resistant)
  • MDAI2 w/ plasmids pCT6 & pET-GFP
    • AI-2 reporter & LuxS null mutant (E. coli, does not produce AI-2, amp and kan-resistant)

Removed these strains from 4°C -- they are stored in soft agar stab cultures Made one streak plate on LB+amp for each strain from stabs

  • placed in 374°C (rm. 1239)

Made one 2mL LB+amp broth cultures in a 15mL tube for each strain from stabs

  • placed in 30°C, 200 rpm shaking (rm. 1230)

Placed agar stabs and LB+amp broth in 4°C

~1.5 hrs work

Created protocol/plan for experiment. Testing AI-2 Response

--Infekt 00:26, 18 July 2010 (UTC)

7/19/10

Alex and Marcus

7/17 plates are no good -- they needed kanamycin

  • plates were discarded
  • broth cultures had kanamycin added a day later -- too late to be cryostored, but they were transfered to ERB 4°C

Made LB agar 

-20 g powder/500 mL DI water
  • autoclaved 30 min (sterilize time) 255°C
  • poured 4 LB plates
  • poured 6 LB+amp plates (100 μg/mL ampicillin)
  • poured 10 LB+amp+kan plates (50 μg/mL kanamycin)
  • all plates left to cool in ERB 1230

Obtained a rotor for 1.5mL tubes for the centrifuge from Rodger Pinto

  • placed in centrifuge in cold room ERB 1224

Obtained LuxS (strain JW2662-1) & MarC (for Jeremy Minty) null mutant E. coli strains from [http://cgsc.biology.yale.edu/ CGSC]

  • retrieved from Lin 4°C
  • made one spread plate on LB for each strain following [protocol]
  • placed both plates in ERB 37°C

Transfered iGEM cryobox from Lin Lab -20°C to ERB -20°C

~6 hrs

--Infekt 23:49, 19 July 2010 (UTC)----

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