Yeast media
Yeast media kdorfman Mon, 02/08/2016 - 15:03284 Yeast Plate Code
284 Yeast Plate Code kdorfman Wed, 08/23/2023 - 15:49MV + Ade
MV + Ade kdorfman Wed, 10/18/2017 - 16:42For Bio 284
- 0.15 g YNB
- 0.52 g ammonium sulfate
- 2.0 g agar
- 82 mL H2O
- 8.0 mL 1 mg/mL adenine
- 10.0 mL 20% glucose (final conc = 2g/100 mL)
Minimal Vitamin Medium plus Adenine
Label plates with "+ ADE"
Make sure it's a plus sign!
Autoclave an appropriately sized graduated cylinder to measure the glucose and adenine solutions.
Mark with two blue lines
*Yeast Nitrogen Base without amino acids and ammonium sulfate
MV
MV kdorfman Mon, 02/08/2016 - 15:04Minimal Vitamin Medium for Bio 284
Autoclave an appropriately sized graduated cylinder to measure the glucose solution.
*Difco Yeast Nitrogen Base w/o Amino Acids and w/o Ammonium Sulfate
Mark with a single blue line
SC -Leu High Ade
SC -Leu High Ade kdorfman Sun, 01/19/2020 - 16:39Plates for Jeff Laney's Cell & Molecular Biology Lab
8X SC-HLT hi Ade concentrate
8X SC-HLT hi Ade concentrate kdorfman Fri, 01/22/2021 - 22:01Concentrate to make SC-HLT High Ade agar
add 125 mL heated concentrate to autoclaved 875 mL water + 20 g agar right out of the autoclave and still hot
Laney's SC high ade
Laney's SC high ade kdorfman Sun, 01/26/2020 - 16:22From Jeff Laney's recipe:
Per liter of medium, mix the following asceptically, and pre heat:
Component | Volume (mL) |
---|---|
10X SC-AHLT | 100 |
10X YNB+AS | 100 |
10X 20% glucose | 100 |
10X Adenine hemi sulfate | 100 |
50X Tryptophan | 20 |
100X Histidine HCl | 10 |
Add to 20g agar in 667.5 mL water, autoclaved 30 minutes
Mark with one green line.
SC -Leu High Ade 4x + powder
SC -Leu High Ade 4x + powder kdorfman Sun, 01/26/2020 - 16:26SC -Leu High Ade 4x + powder
Use the 4x SC-AHLT to mix dry ingredients
(4x already made; not enough dry left to start over.)
per Liter:
Agar 20 g + 620 mL water. Autoclave
Meanwhile, make:
Component | Volume (mL) |
---|---|
4X SC-AHLT | 250 |
10X 20% glucose | 100 |
50X Tryptophan | 20 |
100X Histidine HCl | 10 |
Add dry:
Component | g |
---|---|
YNB | 1.7 |
Adenine hemi sulfate | 0.2 |
Ammonium Sulfate | 5 |
Filter sterilize, warm up, then add to molten agar
Mark with one green line
SC -Leu High Ade 4X
SC -Leu High Ade 4X kdorfman Sun, 01/26/2020 - 16:23Using dilution factor 4:
(Because 10x won't go into solution)
But the agar is so viscous that bubbles don't reach the surface. Hard to pour.
Mark with one green line
SC-L high Ade from concentrate
SC-L high Ade from concentrate kdorfman Fri, 01/22/2021 - 20:55Mark with one green line.
SC-L high Ade mostly powder
SC-L high Ade mostly powder kdorfman Tue, 02/04/2020 - 13:41High concentration agar is difficult to work with.
- Tends to boil over, so needs a lot of water in the autoclave pan
- Hard to get into solution - frequently needs re-autoclaving
- So viscous that bubbles don't rise to the surface
So
- mix the ingredients that can't be autoclaved in as little water as possible
- Only use high concentration stock solutions for micro-ingredients
Per Liter:
dry ingredient | g |
---|---|
SC-AHLT | 1.64 |
YNB | 1.7 |
Ammonium sulfate | 5 |
glucose | 20 |
stock solutions | mL |
---|---|
50X Tryptophan | 20 |
100X Histidine HCl | 10 |
10X Adenine hemi sulfate | 100 |
plus water to 150 mL
filter sterilize
Autoclave
- 20 g agar
- 850 mL water
Mix and pour
SC -Leu-low-Ade
SC -Leu-low-Ade kdorfman Thu, 01/16/2020 - 22:07SC-Leu low-Ade plates for Jeff Laney's cell and molecular biology lab
Laney's original
Laney's original kdorfman Sun, 01/26/2020 - 17:02SC-Leu low-Ade plates
From Jeff Laney's recipe:
(BUT, can't make 10X SC-AHLT)
Per liter of medium, mix the following asceptically, and pre heat:
Component | Volume (mL) |
---|---|
10X SC-AHLT | 100 |
10X YNB+AS | 100 |
10X 20% glucose | 100 |
10X Adenine hemi sulfate | 2.5 |
50X Tryptophan | 20 |
100X Histidine HCl | 10 |
Add to 20g agar in 667.5 mL water, autoclaved 30 minutes
Mark with one green and one blue line.
SC -Leu-low-Ade 4X + powder
SC -Leu-low-Ade 4X + powder kdorfman Sun, 01/26/2020 - 17:02SC -Leu low Ade 4x + powder
Use the 4x SC-AHLT to mix dry ingredients
(4x already made; not enough dry left to start over.)
per Liter:
Agar 20 g + 617.5 mL water. Autoclave
Meanwhile, make:
Component | Volume (mL) |
---|---|
4X SC-AHLT | 250 |
10X Adenine hemi sulfate | 2.5 |
10X 20% glucose | 100 |
50X Tryptophan | 20 |
100X Histidine HCl | 10 |
Add dry:
Component | g |
---|---|
YNB | 1.7 |
Ammonium Sulfate | 5 |
Filter sterilize, warm up, then add to molten agar
Mark with one green and one blue line
SC -Leu-low-Ade 4X
SC -Leu-low-Ade 4X kdorfman Sun, 01/26/2020 - 17:01Using 4X SC-AHLT because 10x won't go into solution:
(BUT: Agar is too viscous - bubbles won't rise to the surface before it sets up.)
Mark with one green and one blue line.
SC-AHLT low Ade mostly powder
SC-AHLT low Ade mostly powder kdorfman Tue, 02/04/2020 - 13:26High concentration agar is difficult to work with.
- Tends to boil over, so needs a lot of water in the autoclave pan
- Hard to get into solution - frequently needs re-autoclaving
- So viscous that bubbles don't rise to the surface
So
- mix the ingredients that can't be autoclaved in as little water as possible
- Only use high concentration stock solutions for micro-ingredients
Per Liter:
dry ingredient | g |
---|---|
SC-AHLT | 1.64 |
YNB | 1.7 |
Ammonium sulfate | 5 |
glucose | 20 |
stock solutions | mL |
---|---|
10X Adenine hemi sulfate | 2.5 |
50X Tryptophan | 20 |
100X Histidine HCl | 10 |
plus water to 150 mL
filter sterilize
Autoclave
- 20 g agar
- 850 mL water
Mix and pour
Mark with one green and one blue line.
SC-AHLT
SC-AHLT kdorfman Wed, 01/15/2020 - 21:45SC minus (adenine, histidine, leucine, tryptophan)
4X will dissolve. Almost completely clear after overnight stirring. Crystal clear after autoclaving.
From the manufacturer:
Commonly added to YNB, nitrogen and glucose, at suggested g/L, for a complete yeast media. Mix with stirring for 10-15 minutes, and filter sterilize or autoclave at 121°C for 15 minutes. To prepare plates, autoclave agar separately and add to sterile medium, or add agar to liquid medium and adjust to pH 5.8-6.0 before autoclaving.
Suggested g/L: 1.64
Storage Temperature: 2-8 C
Jeff's recipe: 10X SC –AHLT 16.4 g/L
Dissolve 16.4 g of Sunrise Science SC –Adenine, –Histidine, –Leucine and –Tryptophan in 1 L distilled water and filter sterilize.
Store in the dark at 4 ˚C.
Will not go into solution.
SC-HLT Ade-free Concentrate
SC-HLT Ade-free Concentrate kdorfman Wed, 02/02/2022 - 19:18Concentrate to make SC-L High Ade or Low Ade agar
Make this concentrate, then add adenine hemi-sulfate.
for low-Ade, add 5 mg Ade per L of concentrate
for high-Ade, add 1.6 g Ade per L of concentrate
to make high-Ade from low-Ade concentrate, add 1.595g Ade per L, or 0.1595g per 100 mL
Then filter sterilize. For every L of medium, combine 20g agar autoclaved in 875 mL water with 125 mL warmed up concentrate.
add 125 mL concentrate to autoclaved 875 mL water + 20 g agar
SC-complete-high-Ade
SC-complete-high-Ade kdorfman Wed, 01/15/2020 - 22:12Liquid Medium
Using 4X SC-AHLT (because 10x won't go into solution):
Bring to final volume with water, then filter sterilize.
Jeff Laney's original (discontinued - because you can't make 10X SC-AHLT) recipe:
Component | Volume (mL) |
---|---|
10X SC–AHLT | 25 |
10X YNB+AS | 25 |
10X SC–AHLT | 25 |
20% Glucose | 25 |
10x adenine hemi sulfate | 25 |
50X Tryptophan | 5 |
100X Histidine | 2.5 |
10X Leucine | 25 |
distilled water | to final volume of 250 |
Filter sterilize
Store at room temperature
SC -Leu High Ade
SC -Leu High Ade kdorfman Sun, 01/19/2020 - 13:33Recipe from Jeff Laney. (But 10X SC-AHLT wouldn't go into solution. See 4X recipe.)
Component | Volume (mL) |
---|---|
10X SC-AHLT | 100 |
10X YNB+AS | 100 |
10X 20% glucose | 100 |
10X Adenine hemi sulfate | 100 |
50X Tryptophan | 20 |
100X Histidine HCl | 10 |
Using a different dilution factor:
Mark with one green line
YEAD
YEAD kdorfman Fri, 08/04/2023 - 22:03Yeast Extract Adenine Dextrose Medium
Ade mutants grow well on this without turning red. Better for keeping the parental strains white longer.
1 gram Yeast Extract 2 grams anhydrous dextrose (glucose) 2 grams Agar 8 mL adenine stock solution (1 mg/mL)
Adenine Stock Solution
400 mg adenine in 400 ml water (1 mg/ml) Store at room temperature. Use 2 ml stock solution for 100 ml of medium; reduce the water added to the medium by 2 ml.
Remember to autoclave an appropriate sized graduated cylinder to measure the glucose solution in the sterile hood.
YED
YED kdorfman Mon, 02/08/2016 - 19:22Yeast Extract Dextrose
For Bio 284
Remember to autoclave an appropriate sized graduated cylinder to measure the glucose solution in the sterile hood.
NOTE: 900 mL may boil over in a 1 L bottle in the autoclave. Try:
- Autoclave:
- YE + agar in 800 mL
- 100 mL H2O
- mix in BSC
- Add 100 mL sterile 20% glucose
- mix and pour
Mark with a single red line.
YEKAC
YEKAC kdorfman Mon, 02/08/2016 - 19:29Sporulation Medium for Bio 284
If you have concentrated potassium acetate solution, then:
- 20.4 mL 5M KOAc per liter
- 12.75 mL 8M KOAc per liter Fisher AAJ63372AE
Mark with a single black line.
If you are using potassium acetate powder, then:
YEPAD
YEPAD kdorfman Mon, 08/07/2023 - 17:05YEPAD medium (Yeast Extract + Peptone + Adenine + Dextrose) contains the same ingredients as YEAD but has peptone added. This is a very rich medium used for storing yeast strains.
1 gram Yeast Extract 2 grams Peptone 2 grams anhydrous dextrose (glucose) 2 grams Agar (agar-agar; gum agar) 8 mL adenine stock solution 92 ml water
YEPAD for freezing
YEPAD for freezing kdorfman Wed, 08/16/2023 - 18:06Yeast strains, transformed or untransformed, can be maintained as colonies on solid media at 4° and restreaking every 2 to 4 weeks. Alternatively, strains may be stored at -80 indefinitely. This is preferable in that it reduces the likelihood of accumulating spontaneous mutations.
To make frozen stocks of yeast strains: -Use sterile technique and sterile solutions throughout this method.-
Grow a starter culture at 30 with shaking (250 rpm) until it reaches saturation.
In a 1.8 ml cryotube, mix 0.5 ml of the saturated culture with 0.5 ml of YPD containing 20% glycerol.
Flash freeze the tube in liquid nitrogen and store at -80.
To use, chip out a few pieces of the frozen stock using a sterile pipette tip or sterile toothpick and streak onto a plate containing the appropriate solid media.
Allow the yeast to grow at 30 until colonies appear (2-6 days).
YNB + AS
YNB + AS kdorfman Wed, 01/15/2020 - 21:4410X YNB+AS
17 g/L YNB and 50 g/L AS
Dissolve 17 g Difco Yeast Nitrogen Base w/o Amino Acids and w/o Ammonium Sulfate and 50 g Ammonium Sulfate in 1L distilled water.
Autoclave for 15 min.
Store at room temp.
YPD
YPD kdorfman Mon, 02/08/2016 - 15:04Yeast extract-Peptone-Dextrose medium: