Pichia Adenine Dropout Agar, PAD Agar | 250g
- Product Code: GMNB-PAD124
- Availability: In Stock
Tags: PAD Agar
Pichia Adenine Dropout (PAD) Agar, 250 g
Production
This media is prepared in small batches and will be prepared
after the purchase, taking around 5 to 10 business days for handling.
Application
For selecting ADE2 gene-deficient Pichia Pastoris colonies.
Why use PAD
rather than minimal media for ADE-deficient Pichia selection?
Although minimal media (MY), minimal media with
glycerol/methanol (MGY/MM) and complex media (CY) can also be used to select
ADE mutant colonies. However, MY and even CY are much slower and less efficient
at picking out white/red colonies compared to PAD media because Pichia growth
on MY is slow (small colonies are difficult to compare visually) and CY
contains yeast extract, and yeast extract contains adenine, so all colonies
initially appear the same white, making it difficult to achieve high-resolution
colour assays (the contrast between pure white/white/pink/red) between
colonies. Besides, MY & CY are a bit complex to prepare because of the
complex trace elements stock & vitamins stock need to be prepared
separately.
Components,
per litre
Mixture A |
|
Yeast Nitrogen
Base w/o amino acids and ammonium sulfate |
3.4 g |
Dextrose |
20 g |
(NH4)2SO4 |
10 g |
Mixture B |
|
Uracil |
20 mg |
L-Tryptophan |
20 mg |
L-Histidine |
20 mg |
L-Arginine |
20 mg |
L-Methionine |
20 mg |
L-Tyrosine |
30 mg |
L-Leucine |
30 mg |
L-Isoleucine |
30 mg |
L-Lysine |
30 mg |
L-Phenylalanine |
50 mg |
L-Glutamic
acid |
100 mg |
L-Aspartic acid |
100 mg |
L-Valine |
150 mg |
L-Threonine |
200 mg |
L-Serine |
400 mg |
Preparation
[Standard preparation, recommended]
1. Prepare 0.5 litre of 10X SD stock solution: Dissolve 167 g
mixture A in distilled or Milli-Q water and mix well, make it to 500 ml. Sterilise by 0.22µm filtration (recommended)
or autoclave at 115℃ for 30 minutes.
Store the stock at room temperature. The shelf life of this solution is
approximately one year.
2. Weigh 1.22 g of mixture B, and a suitable amount of agar (1~2% w/v recommended), in distilled or Milli-Q Water, make it to 900ml. Sterilise by autoclaving at 121°C for 15 minutes.
3. Add 100 mL of sterile 10X SD stock into the freshly autoclaved
(no longer boiling) mixture B solution, mix well and pour the plates while
hot. Store the PAD agar plates at 4°C.
The shelf life is several months.
[Fast preparation, may cause a dark colour of the agar]
1. Weigh 33.4 g of mixture A (SD medium), 1.22 g of mixture B, and a suitable amount of Agar [1] in distilled or Milli-Q Water, make it to 1000ml.
2. Sterilise by autoclaving at 121℃ for 15 minutes [2]. Aliquot as needed.
[1]
1~2% w/v recommended
[2]
If equipment allows, autoclave at
115℃ for 30 minutes to prevent the darkening of the solution due to a Maillard
reaction on glucose catalysed by ammonium sulphate and heat.