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Team:Paris Bettencourt/Notebook/Drug Screening/Wednesday 24th July.html
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| - | <h3> | + | <h3>Wednesday 24<sup>th</sup> July</h3> |
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Latest revision as of 13:06, 13 August 2013
Drug Screening
ASDFWednesday 24th July
Place your twit here
Made 500 ml of M9 Minimal Plates with Sulfur Dropout Powder accouding to the following recipe:
Recipe: M9 Minimal Plates with Sulfur Dropout Powder
These stock solutions can be pre-sterilized and stored indefinitely:
5x M9 Salts (from scratch)
|
Reagent |
Quantity |
|
Na2HPO4 |
17 g |
|
KH2PO4 |
7.5 g |
|
NaCl |
1.25 g |
|
(NH4)2SO4 |
3.0 g |
Bring to a total volume of 500 ml.
Adjust pH to 7.4 with NaOH.
Sterilize by autoclaving or filtration
or
5x M9 Salts (from Difco Powder)
|
Reagent |
Quantity |
|
M9 Salts |
28.2 g |
Bring to a total volume of 500 mL.
Sterilize by autoclaving or filtration
2x Yeast Plate Agar
|
Reagent |
Quantity |
|
Agar |
30 g |
Bring to a total volume of 500 ml.
Sterilize by autoclaving
5x Amino Acid Dropout Powder
|
Reagent |
Quantity |
|
Sulfur dropout powder |
0.5 g |
Bring to a total volume of 200 ml.
Sterilize by autoclaving
Sulfur dropout powder is a rich supplement mix with cysteine and methionine omitted:
|
Amino acids |
|
Nucleotide bases |
|
|
|
|
Alanine |
2.0 g |
Leucine |
10.0 g |
Adenine |
0.5 g |
|
Arginine |
2.0 g |
Lysine |
2.0 g |
Uracil |
2.0 g |
|
Asparagine |
2.0 g |
Methionine |
0.0 g |
|
|
|
Aspartic acid |
2.0 g |
Phenylalanine |
2.0 g |
Vitamins |
|
|
Cysteine |
0.0 g |
Proline |
2.0 g |
p-Aminobenzoic acid |
0.2 g |
|
Glutamic acid |
2.0 g |
Serine |
2.0 g |
Inositol |
2.0 g |
|
Histidine |
2.0 g |
Tyrosine |
2.0 g |
|
|
|
Isoleucine |
2.0 g |
Valine |
2.0 g |
|
|
Recipe: M9 Minimal Plates with Sulfur Dropout Powder
The plates can be made fresh from pre-sterilized stock solutions.
|
Reagent |
Quantity |
|
5x M9 Salts |
200 mi |
|
5x Amino Acid Dropout Powder |
200 mi |
|
1 M MgSO4 |
2 ml |
|
20% Glucose |
20 ml |
|
1M CaCl2 |
0.1 ml |
Bring to a total volume of 500 mL
Stir and warm on a hot plate to about 60 C (hot to the touch)
Melt 500 ml 2x Yeast Plate Agar in the microwave for ~8 minutes on high
Cool to about 60 C (hot to the touch)
This takes about 30 minutes on the bench or 5 minutes in a water bath.
Add the 2x Yeast Plate agar to the other reagents to make 1 L of media.
This recipe will prepare 20 plates at 25 ml / plate.
made 7 plates without antibiotics and 20 with Chloramphenicol and Spectinomycin - by mistake.
Used one plate without AB to plate the following strains:
sD007=sD001+pD005
sD008=sD001+pD006
sD013= sD001+pD005+ pD004
sD015= sD001+pD006+ pD004
