Team:Michigan/Media Recipes

From 2010.igem.org

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'''LB media'''
'''LB media'''
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*10 g LB broth
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*10 g LB medium
*500 mL DI water
*500 mL DI water
''autoclave to sterilize''
''autoclave to sterilize''
'''LB media with agar'''
'''LB media with agar'''
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*10 g LB broth
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*10 g LB medium
*7.5 g agar
*7.5 g agar
*500 mL DI water
*500 mL DI water
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'''10mM dNTP mix for PCR'''
'''10mM dNTP mix for PCR'''
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''MAKE SURE ALL REAGENTS ARE WELL MIXED BEFORE STARTING BY TOUCH CENTRIFUGING OR FLICKING''
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''MAKE SURE ALL REAGENTS ARE WELL MIXED BEFORE STARTING BY PULSE CENTRIFUGING OR FLICKING''
*100 uL of 100mM dATP
*100 uL of 100mM dATP
*100 uL of 100mM dTTP
*100 uL of 100mM dTTP
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'''1.5 mg/mL Cyclohexanecarboxylic acid solution'''
'''1.5 mg/mL Cyclohexanecarboxylic acid solution'''
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Cyclohexanecarboxylic acid is volatile should only be worked with in a hood.  For accurate measurement use covered scale with sliding doors in the hood. Stored in the corrosive cabinet
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Cyclohexanecarboxylic acid is volatile and should only be used under a hood.  For accurate measurement, use a covered scale with sliding doors (in the hood). Store in the corrosive cabinet.
*0.075 g cyclohexanecarboxylic acid
*0.075 g cyclohexanecarboxylic acid
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*0.002 g of FeCl3
*0.002 g of FeCl3
*318 mL of DI H2O  
*318 mL of DI H2O  
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Autoclave at 15 psi and 121C for 30 minutes
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''Autoclave at 15 psi and 121°C for 30 minutes''
After the media has cooled completely add:
After the media has cooled completely add:
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Scale up the amount of  0.1 M NaOH solution to add to the remaining volume of media (approaching final amount slowly)
Scale up the amount of  0.1 M NaOH solution to add to the remaining volume of media (approaching final amount slowly)
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On 8/3/2010 approximately 20 uL of 0.1 M NaOH had to be added to 1 mL to get to a pH between 8 and 9 (tested with pH paper) and 675 uL of 0.1 M NaOH was added into the 45 mL of remaining media.
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On 8/3/2010, approximately 20 uL of 0.1 M NaOH had to be added to 1 mL to get to a pH between 8 and 9 (tested with pH paper) and 675 uL of 0.1 M NaOH was added into the 45 mL of remaining media.
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Latest revision as of 23:52, 26 October 2010


Michigan Header





Media Recipes

General Recipes

Oil Sands Recipes

General Recipes

LB media

  • 10 g LB medium
  • 500 mL DI water

autoclave to sterilize

LB media with agar

  • 10 g LB medium
  • 7.5 g agar
  • 500 mL DI water

autoclave to sterilize

add antibiotics after autoclaving when warm to touch (about 50°C)

pour plates when warm to the touch (about 50°C)

0.1 M CaCl2 solution

  • 0.735 g of CaCl2*2 H20
  • 50 mL of DI water

filter sterilize

1.0 M CaCl2 solution

  • 7.35 g of CaCl2*2 H20
  • 50 mL of DI water

filter sterilize

1.0 M MgSO4

  • 12.323 g of MgSO4*7 H20
  • 50 mL of DI water

filter sterilize

Ampicillin Antibiotic Stock Solution (100 mg/mL)

  • 1 g of ampicillin
  • 5 mL of DI water
  • 5 mL of ethanol

filter sterilize and make 1 mL aliquotes to store in the -20°C freezer

Kanamycin Antibiotic Stock Solution (50 mg/mL)

  • 250 mg of kanamycin
  • 5 mL of DI water

filter sterilize and make 1 mL aliquots to store in the -20°C freezer

Tetracycline Antibiotic Stock Solution (50 mg/mL)

  • 250 mg of tetracycline
  • 2.5 mL of DI water
  • 2.5 mL of ethanol

filter sterilize and make 1 mL aliquots to store in the -20°C freezer

IPTG stock solution (1 M)

  • 0.25 g of IPTG
  • 1049 uL of DI water

filter sterilize with .22 um filter

40% glucose solution

  • 20 g of glucose
  • 50 mL of DI water

filter sterilize

0.1 M NaOH solution

BE VERY CAREFUL WHEN MIXING! SOLID NaOH BURNS YOUR SKIN BUT YOU MAY NOT FEEL IT AT FIRST! MAKE SURE ANY SPILLS ARE CLEANED UP VERY WELL!

  • 0.2 g of NaOH
  • 50 mL of sterile water

Filter Sterilize

10mM dNTP mix for PCR

MAKE SURE ALL REAGENTS ARE WELL MIXED BEFORE STARTING BY PULSE CENTRIFUGING OR FLICKING

  • 100 uL of 100mM dATP
  • 100 uL of 100mM dTTP
  • 100 uL of 100mM dGTP
  • 100 uL of 100mM dCTP
  • 600 uL of ultra pure water

aliquot into 50 uL volumes (this will take 20 1.5mL eppendorf tubes)

Oil Sands Recipes

1.5 mg/mL Cyclohexanecarboxylic acid solution

Cyclohexanecarboxylic acid is volatile and should only be used under a hood. For accurate measurement, use a covered scale with sliding doors (in the hood). Store in the corrosive cabinet.

  • 0.075 g cyclohexanecarboxylic acid
  • 50 mL of DI water

filter sterilize

Bushnell-Haas Salts (w/out carbon source)

  • 0.4 g of KH2PO4
  • 0.4 g of K2HPO4
  • 0.4 g of (NH4)2 SO4 or NH4NO3
  • 0.002 g of FeCl3
  • 318 mL of DI H2O

Autoclave at 15 psi and 121°C for 30 minutes

After the media has cooled completely add:

  • 665 uL of 1 M MgSO4 solution
  • 72 uL of 1 M CaCl2 solution

Bushnell-Haas Media with glucose

  • 39.75 mL of Bushnell-Haas Salts
  • 2 mL of 40% glucose solution
  • 8.25 mL of sterile DI water

Bushnell-Haas Media with cyclohexanecarboxylic acid

  • 39.75 mL of Bushnell-Haas Salts
  • 4 mL of cyclohexanecarboxylic acid solution
  • 6.25 mL of sterile DI water

20 mg/mL Casamino Acid Solution

  • 0.18 g Casamino acid
  • 9 mL of DI water

Filter sterilize

Bushnell-Haas Media with glucose and casamino acids

  • 39.75 mL of Bushnell-Haas Salts
  • 2 mL of 40% glucose solution
  • 2 mL of sterile DI water
  • 6.25 mL of casamino acid solution

Bushnell-Haas Media with cyclohexanecarboxylic acid adjusted to pH of 9

  • 39.75 mL of Bushnell-Haas Salts
  • 4 mL of cyclohexanecarboxylic acid solution
  • 6.25 mL of sterile DI water

Test how much 0.1 M NaOH must be added to 1 mL of media to raise it to a pH of 9 (add 1 uL of base at a time)

Scale up the amount of 0.1 M NaOH solution to add to the remaining volume of media (approaching final amount slowly)

On 8/3/2010, approximately 20 uL of 0.1 M NaOH had to be added to 1 mL to get to a pH between 8 and 9 (tested with pH paper) and 675 uL of 0.1 M NaOH was added into the 45 mL of remaining media.

Lab01.png

Lab02.png

Lab04.png

Lab05.png

Lab06.png

Lab09.png

Lab07.png

Lab08.png

Lab10.png

Lab11.png

Retrieved from "http://2010.igem.org/Team:Michigan/Media_Recipes"