TEWLAB Yeast Media – General Description
WHAT IS IN YEAST MEDIA
Yeast media MUST contain the following components for the yeast to grow:
1) A carbon source.
2) A nitrogen source, also called a nutrient base.
In addition, yeast media MAY contain:
1) Various additional nutritional supplements.
2) Various additional "additives", typically drugs or antibiotics.
3) 2% agar for solid medium (plates). Liquid medium is identical, but without agar.
TYPES OF CARBON SOURCES
1) 2% Dextrose, which is the same thing as Glucose. A sugar. Typically abbreviated as
"D" or "Dex" or "Glu". This is by far the most common carbon source, and is preferred
by the yeast.
2) 2% Galactose (Gal). Another sugar, typically used for regulated expression of genes.
3) 2% Raffinose (Raf). Yet another sugar. This sugar is non-repressing (i.e. cells behave as
if glucose is not present), but does not induce Gal genes.
4) 1% Potassium Acetate. Acetate requires respiratory growth. Most commonly used in
sporulation medium.
5) 3% Glycerol. Glycerol requires respiratory growth. Most commonly used to verify that
yeast are not petite (i.e. that they have mitochondrial function).
Note that carbon sources may be combined. There are also others that are not listed – those
above are the most common.
TYPES OF NITROGEN SOURCES/NUTRIENT BASES
1) 1% Yeast Extract + 2% Peptone (YP). This is a complex nutrient base derived from
killed yeast cells. It provides everything yeast need to grow except a carbon source
(although it is typically low in adenine), and so makes non-selective rich medium.
2) Yeast Nitrogen Base with Ammonium Sulfate (YNB + AS). This is a combination of
various ingredients that we purchase as a mix from Difco. It is made from purified
chemicals, and so is used to make selective media.
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TYPES OF NUTRITIONAL SUPPLEMENTS
1) Individual nutrients. These can be added to media individually, typically from 100X
stocks. Doing this is called an "add back". The typical nutrients we use are:
a) Adenine (Ade, A, 1X = 40 ug/ml. NOTE: for this nutrient only, the CSM mix
below does not contain 1X adenine, but instead has 10 ug/ml = 0.25X adenine.
This is done so that ade2 yeast turn red on SD plates.)
b) Arginine (Arg, R, 1X = 50 ug/ml)
c) Histidine (His, H, 1X = 20 ug/ml)
d) Leucine (Leu, L, 1X = 100 ug/ml)
e) Lysine (Lys, K, 1X = 50 ug/ml)
f) Methionine (Met, M, 1X = 20 ug/ml)
g) Tryptophan (Trp, W, 1X = 50 ug/ml)
h) Uracil (Ura, U, 1X = 20 ug/ml)
2) Complete Supplement Mixture (CSM). This is a mixture that we buy from
Bio101/QBiogene that has all of the above nutrients in addition to those listed below,
mixed together in a powder in the right ratio. Media that contain this are thus nonselective (nearly all yeast will have everything they need to grow).
i) Aspartate (Asp, D, 1X = 80 ug/ml)
j) Isoleucine (Ile, I, 1X = 50 ug/ml)
k) Phenylalanine (Phe, F, 1X = 50 ug/ml)
l) Threonine (Thr, T, 1X = 100 ug/ml)
m) Tyrosine (Tyr, Y, 1X = 50 ug/ml)
n) Valine (Val, V, 1X = 140 ug/ml)
3) CSM – some nutrient(s). These are a whole series of other mixes that are derived from
CSM, but from which certain nutrients have been omitted. Such media is thus called a
"dropout" (it has everything except the nutrient that has been dropped out). For example,
CSM - Ura has everything except uracil.
TYPES OF ADDITIVES
1) G418. 200 ug/ml. A drug that kills yeast that don't have the bacterial kanMX4 gene in
their genome.
2) Canavanine. 50 ug/ml. An arginine analog that kills by being incorporated into proteins.
The CAN1 gene encodes an arginine permease that is required for cells to take up
canavanine. Thus, can1 mutant cells are canavanine resistant, CAN1 are sensitive. To
score for canavanine sensitivity, it is critical that the media be free of arginine, as this will
out-compete the canavanine for the Can1 permease. This means that only Arg+ cells can
be selected on canavanine, since they must be able to make their own arginine.
3) 5-FOA. 0.5 g/500 ml. This drug is metabolized to a toxic product by yeast that have a
wild-type URA3 gene. Thus, only ura3 yeast grow on FOA plates. These plates require
special handling when pouring.
4) Many others. There are a wide variety of other drugs and DNA damaging agents we add
to plates. The typical thing is to add them to the medium after autoclaving.
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TYPES OF YEAST MEDIA
There are two principal types of yeast media that are made from the above components – the key
difference between them is the nitrogen source/nutrient base:
1) YPAD. This is rich and non-selective medium. All yeast will grow. Its components are:
a) nutrient base = 1% Yeast Extract + 2% Peptone (YP)
b) supplement = 1X (40 ug/ml) adenine (A)
c) carbon source = 2% Dextrose (D)
There are many variations on the above!
a) YPD, which is the same but without the additional adenine. Yeast will still all
grow, but ade2 yeast will turn red on YPD.
b) YPA with another carbon source. This is again a rich medium, but with a
different carbon source substituted for the Dextrose. The most common are
YPAGal and YPAGlycerol (YPAG).
c) YPAD with an additive. Many drugs can simply be added to YPAD. The most
common for us is YPAD + 200 ug/ml G418.
2) Synthetic Defined (SD). This medium is chemically defined. Synthetic defined medium
contains the following components.
a) nutrient base = YNB + AS
b) a carbon source
c) a supplement mix (optional).
As with rich medium, there are many variations on the above!
a) When the carbon source = Dextrose and there is NO supplement mix, the above
makes SD Glu, which we more commonly call "minimal" medium. Only wildtype (i.e. prototrophic) yeast will grow, since no nutritional supplements have
been added.
b) When the carbon source = Dextrose and the supplement mix = CSM, the plate
now contains everything that yeast need to grow, so the medium is non-selective.
However, it is still synthetically defined. This is SD Glu complete, also called SC
Glu (for Synthetic Complete). We commonly refer to this simply as "complete".
c) The most common variation is carbon source = Dextrose and supplement mix =
CSM – some nutrient(s), so called drop-out media. Thus, only strains
prototrophic for dropped out nutrient(s) will grow, which forms the basis of the
most common selections. These plates can be called, for example, SD Glu – Ura,
SC Glu – Ura, Glu – Ura, or simply –Ura (note that if the carbon source is not
specified, you typically assume it is glucose).
d) Beyond this, one can of course vary the carbon source, add all sorts of additives,
additional adenine supplement, etc.
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