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