Yeast media

Yeast media kdorfman Mon, 02/08/2016 - 15:03
284 Yeast Plate Code.docx

MV + Ade

MV + Ade kdorfman Wed, 10/18/2017 - 16:42
  • 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.

To make MV+ADE agar plates
pour mm plate diameter
pour mL per plate
start with: mL water
add: g YNB*, stir till dissolved
add: g ammonium sulfate, stir till dissolved
add: g agar, leave stir bar in
autoclave for: minutes
add asceptically mL filter sterilized 20% glucose
add asceptically: mL filter-sterilized adenine (1mg/mL)
for a final volume of mL

Mark with two blue lines

*Yeast Nitrogen Base without amino acids and ammonium sulfate

MV

MV kdorfman Mon, 02/08/2016 - 15:04

Minimal Vitamin Medium

Autoclave an appropriately sized graduated cylinder to measure the glucose solution.

To make MV agar plates
pour mm plate diameter
pour mL per plate
start with: mL water
add: g YNB*, stir till dissolved
add: g ammonium sulfate, stir till dissolved
add: g agar, leave stir bar in
autoclave for: minutes
add asceptically mL filter sterilized 20% glucose
for a final volume of mL

*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 - 16:39

Plates 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 - 22:01

Concentrate 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

to make enough concentrate for:
Liter(s) SC-HLT high Ade agar medium
start with
mL water
add:
g YNB
and:
g ammonium sulfate
and:
g SC -HLT
autoclave for:
minutes (to get it into solution)
while it's still hot, add:
g glucose
and:
g Adenine hemi sulfate
and:
g tryptophane
and:
mL his 100X concentrate.
Bring to a final volume of:
mL 8X SC-L high Ade and filter sterilize

Laney's SC high ade

Laney's SC high ade kdorfman Sun, 01/26/2020 - 16:22

From 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 - 16:26

SC -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 - 16:23

Using 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.

To make
SC-leu high ade agar plates
with mm plate diameter
pour mL per plate
start with: mL water
add: g agar, leave stir bar in
autoclave for: minutes
Meanwhile, using X SC-AHLT
Asceptically mix together,
warm up, then add to autoclaved agar:		 mL SC-AHLT,
and : mL 10x YNB+AS ,
and: mL 20% glucose ,
and: mL 10X Adenine hemi sulfate,
and: mL 50X Tryptophan,
and: mL 100X Histidine HCl.
for a final volume of mL

Mark with one green line

SC-L high Ade from concentrate

SC-L high Ade from concentrate kdorfman Fri, 01/22/2021 - 20:55
To make SC -L high Ade plates (100 mm)
mix: g agar
into: mL water
autoclave for: minutes
while it's still hot, asceptically add: mL heated 8X SC-L high Ade concentrate
for a final volume of: mL. Mix thoroughly before pouring plates

Mark with one green line.

SC-L high Ade mostly powder

SC-L high Ade mostly powder kdorfman Tue, 02/04/2020 - 13:41

High 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 - 22:07

SC-Leu low-Ade plates for Jeff Laney's cell and molecular biology lab

Laney's original

Laney's original kdorfman Sun, 01/26/2020 - 17:02

SC-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 - 17:02

SC -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 - 17:01

Using 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.)

To make SC-leu low ade agar plates
with mm plate diameter
pour mL per plate
start with: mL water
add: g agar, leave stir bar in
autoclave for: minutes
Meanwhile, using X SC-AHLT
Asceptically mix together,
warm up, then add to autoclaved agar:		 mL SC-AHLT,
and : mL 10X YNB+AS,
and: mL 20% Glucose,
and: mL 10x adenine hemi sulfate,
and: mL 50X Tryptophan
and: mL 100X Histidine .
for a final volume of mL

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 - 13:26

High 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 - 21:45

SC minus (adenine, histidine, leucine, tryptophan)

Sunrise Science 1330-030

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 - 19:18

Concentrate 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

to make enough concentrate for:
Liter(s) SC-L high Ade or SC-L low Ade agar medium
start with
mL water
add:
g YNB
and:
g ammonium sulfate
and:
g SC -HLT
autoclave for:
minutes (to get it into solution)
while it's still hot, add:
g glucose
and:
g tryptophane
and:
mL his 100X concentrate.
Bring to a final volume of:
mL 8X SC-HLT Ade-free. Add the appropriate amount of Ade and filter sterilize

SC-complete-high-Ade

SC-complete-high-Ade kdorfman Wed, 01/15/2020 - 22:12

Liquid Medium

Using 4X SC-AHLT (because 10x won't go into solution):

To make mL SC-complete high ade liquid medium
Meanwhile, using X SC-AHLT
Mix: mL 10X SC–AHLT,
and : mL 10X YNB+AS,
and: mL 20% Glucose,
and: mL 10x adenine hemi sulfate,
and: mL 50X Tryptophan ,
and: mL 100X Histidine .
and: mL 10X Leucine

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 - 13:33

Recipe 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:

To make
SC-leu high ade agar plates
with mm plate diameter
pour mL per plate
start with: mL water
add: g agar, leave stir bar in
autoclave for: minutes
Meanwhile, using X SC-AHLT
Asceptically mix together,
warm up, then add to autoclaved agar:		 mL SC-AHLT,
and : mL 10x YNB+AS ,
and: mL 20% glucose ,
and: mL 10X Adenine hemi sulfate,
and: mL 50X Tryptophan,
and: mL 100X Histidine HCl.
for a final volume of mL

Mark with one green line

YED

YED kdorfman Mon, 02/08/2016 - 19:22

Yeast Extract Dextrose

Remember to autoclave an appropriate sized graduated cylinder to measure the glucose solution in the sterile hood.

To make YED agar plates
at mm plate diameter
pour mL per plate
start with: mL water
add: g yeast extract, stir till dissolved
add: g agar, leave stir bar in
autoclave for: minutes
add asceptically mL filter sterilized 20% glucose
for a final volume of mL

Mark with a single red line.

YEKAC

YEKAC kdorfman Mon, 02/08/2016 - 19:29

Sporulation Medium

To make YEKAC agar plates
at mm plate diameter
pour mL per plate
start with: mL water
add: g potassium acetate, stir till dissolved
add: g yeast extract, stir till dissolved
add: g agar, leave stir bar in
autoclave for: minutes

OR

  • 20.4 mL 5M KOAc per liter
  • 12.75 mL 8M KOAc per liter Fisher AAJ63372AE

Mark with a single black line.

YNB + AS

YNB + AS kdorfman Wed, 01/15/2020 - 21:44

10X 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 - 15:04

Yeast extract-Peptone-Dextrose medium:

To make YPD agar plates
at mm plate diameter
pour mL per plate
start with: mL water
add: g peptone broth, stir till dissolved
add: g yeast extract, stir till dissolved
add: g agar, leave stir bar in
autoclave for: minutes
add asceptically mL filter sterilized 20% glucose
for a final volume of mL YPD agar