Craig’s Media

Chlamy Growth Media

Adapted from various in-lab sources

Stock Solutions

Component % chemical in stock in 1L stock
#1 Trace Elements Look Right –
#2 Na Citrate·2H2O 10% 100g
#3 FeCl3·6H2O 1% 10g
#4 CaCl2·6H2O 5.3% 53
#5 MgSO4·7H2O 10% 100g
#6 NH4NO3 10% 100g
#7 KH2PO4 10% 100g
#8 K2HPO4 10% 100g
Trace Elements
Component in 1L stock
H3BO3 1g
ZnSO4·7H2O 1g
MnSO4·H2O 0.303g
CoCl2·6H2O 0.2g
Na2MoO4·2H2O 0.2g
CuSO4 0.04g

Recipes

For 1L of media

Component M M/NO3 M – N R R/NO3 R – N SG II SG II/NO3
#1 1 ml 1 ml 1 ml 1 ml 1 ml 1 ml 1 ml 1 ml
#2 5 ml 5 ml 5 ml 5 ml 5 ml 5 ml 5 ml 5 ml
#3 1 ml 1 ml 1 ml 1 ml 1 ml 1 ml 1 ml 1 ml
#4 1 ml 1 ml 1 ml 1 ml 1 ml 1 ml 1 ml 1 ml
#5 3 ml 3 ml 3 ml 3 ml 3 ml 3 ml 3 ml 3 ml
#6 3 ml – – 4 ml – – 3 ml 3 ml
#7 1 ml 0.8 ml – 2 ml 1 ml – – –
#8 1 ml 1.5 ml 2 ml 3.5 ml 1.75 ml 3.5 ml – –
1M KNO3 – 4 ml – – 4 ml – – 4 ml
2.2M Na Acetate – – – 6 ml 6 ml 6 ml – –
K2HPO4 – – – – – – 1.5 g 1.5 g
NaH2PO4·H2O – – – – – – 3.67 g 3.67 g
Na Acetate·3H2O – – – – – – 2 g 2 g
For arginine requiring mutants add 2ml of 10% arginine to 1L of media.

TAP Media

Recipe

Stock Solution For 1 L
1M Tris base 20 ml
Phosphate Buffer II 1 ml
Hutner’s Trace Metals 1 ml
Solution A 10 ml
Glacial acetic acid
(pH to 7.0)		 1 ml

Stock Solutions

Phosphate Buffer II
Component For 100 ml
K2HPO4 10.8 g
KH2PO4 5.6 g
Solution A
Component for 500 ml
NH4Cl 20 g
MgSO4·7H2O 5 g
CaCl2·2H2O 2.5 g

Hutner’s Trace Metals

  1. Dissolve 50g of acid free EDTA in 250 ml of ddH2O. Heat to dissolve.
  2. Dissolve the following, one by one, in order, heating to approximately 100° in 550 ml ddH2O.
    BO3H3 11.4 g
    ZnSO4·7H2O 22.0 g
    MnCl2·4H2O 5.06 g
    FeSO4·7H2O 4.99 g
    CoCl2·6H2O 1.61 g
    CuSO4·5H2O 1.57 g
    Mo7O24(NH4)6·4H2O
    (Ammonium Molybdate)    		 1.1 g
  3. Mix the two solutions together. The resulting solution should be blue-green.
  4. Heat to 100°. Cool slightly, but don’t let the temperature drop below 80°-90°.
  5. Adjust pH to 6.5 to 6.8 with 20% KOH (approximately 83 ml). Don’t let the temperature drop below 70 ° until after the pH is adjusted.
  6. Make up to 1L and let stand in a 2L Erlenmeyer flask, stoppered loosly. The colour should change from green to purple.
  7. Remove the rust-coloured precipitate by filtering, with suction, through 3 layers of Whatman #1 filter paper in a Buchner funnel. Repeat until no more precipitate is seen on the filter paper.
  8. Store in a brown bottle at 4°.

Gorman, D.S. and R.P. Levine (1965). Proc. Natl. Acad. Sci. USA 54:1665-1669.
Hutner, et al. (1950). Methods in Enzymology, 23:68.
Sager, R. and S. Granick (1953). Ann. N.Y. Acad. Sci. 56:831-838.

This page is, as always, Copyright (c) 1998 by Craig D. Amundsen. All rights reserved.