Methods

Following are recipes for all standard media prepared by the Media Preparation Facility.

3% Agar

  • 7.5g Bacto Agar/ 250 ml H2O

4% Agar

  • 10.0g Bacto Agar/ 250 ml H2O

40% Glucose

  • 400 g/l Glucose
  • autoclave

 

LB Broth, Miller

  • 10.0 g/l Difco Bacto Tryptone
  • 5.0 g/l Difco Yeast Extrct
  • 10.0 g/l NaCl
  • 10.0 ml/l 1.0 M Tris pH 7.5
  • autoclave

2X LB Broth, Miller

  • double above g/l
  • autoclave

1.5% Agar/LB

  • 6.0 g Bacto Agar/ 400 ml LB

PBS

  • 8.0 g/l NaCl
  • 0.2 g/l KCl
  • 1.15 g/l Na2HPO4
  • 0.2 g/l KH2PO4
  • adjust pH to 7.2
  • autoclave

 

2X S

  • 13.4 g/l Difco Bacto Yeast Nitrogen Base w/o amino acids
  • autoclave

2X S w/o ura, trp

  • 13.4 g/l Difco Bacto Yeast Nitrogen Base w/o amino acids
  • 4.0 g/l amino acid powder w/o ura, trp (prepared by Amon Lab)
  • autoclave

2X S w/o his, leu

  • 13.4 g/l Difco Bacto Yeast Nitrogen Base w/o amino acids
  • 4.0 g/l amino acid powder w/o his, leu (prepared by Amon Lab)

 

Terrific Broth

Solution 1 : Prepared by the Media Preparation Facility

  • 12.0 g/l Difco Bacto Tryptone
  • 24.0 g/l Difco Bacto Yeast Extract or EMD Yeast Extract
  • 4.0 ml/l Glycerol
  • 900 ml H2O
  • autoclave

Solution 2: Prepared by the individual labs

  • 23.1 g/l KH2PO4
  • 125.4 g/l K2HPO4
  • autoclave and cool
  • add 100 ml Solution 2 to 900 ml Solution 1

Versene

  • 8.0 g/l NaCl
  • 0.2 g/l KCl
  • 1.15 g/l Na2HPO4
  • 0.2 g/l KH2PO4
  • 0.2 g/l Na-EDTA
  • 0.015 g/l Phenol red
  • adjust pH to 7.2
  • autoclave

1.05X YEP

  • 10.5 g/l EMD Yeast Extract or Difco Bacto Yeast Extract
  • 21.1 g/l Difco Bacto Peptone
  • autoclave

2.22X YEP

  • 22.2 g/l EMD Yeast Extract or Difco Bacto Yeast Extract
  • 44.4 g/l Difco Bacto Peptone
  • autoclave

 

Trypsin/EDTA

  • 0.20 g/l EDTA
  • 8.00 g/l NaCl
  • 0.40 g/l KCl
  • 0.35 g/l NaHCO3
  • 1.00 g/l Glucose
  • 0.04 g/l Trypsin ( 428 units/ml) (Sigma # T-4665)
  • 0.0075 g/l Phenol red
  • adjust pH to 7.0
  • sterile filter (0.2 um)
  • Freeze immediately after filtering