Describe, in detail, to a freshman undergraduate how to make 1 liter of LB + Kan (100 μg/ml final concentration) + Amp (50 μg /ml final concentration) liquid media. [Include things like how many grams of each component that you use, how much antibiotic (in ml) to add (stock solutions – 100 mg/ml ampicillin, 25 mg/ml kanamycin), and in what type of container you perform the sterilization step.] Show your calculations.

Explanation:

The objective here is to prepare  1 liter of LB + Kan (100 μg/ml final concentration) + Amp (50 μg /ml final concentration) liquid media.

Given that :

the Stock concentration of Amp: 100 mg/ml (since 1 mg/ml = 1000 μg/ml)

it is required that we convert stock concentration in  μg/ml since 100 mg/ml = 100000  μg/ml

However, using formula C₁V₁=C₂V₂ (Ampicilin),

where:

C₁ = 100000 μg/ml,

V₁=?,

C₂= 50  μg/ml,

V₂=1000 ml

100000 μg/ml × V₁ = 50  μg/ml × 1000 ml

V₁ =  50  μg/ml × 1000 ml/100000 μg/ml

V₁ =   0.5 ml

Given that:

the Stock concentration of Kan: 25 mg/ml (since 1 mg/ml = 1000 μg/ml)

it is required that we convert stock concentration in  μg/ml , 25 mg/ml = 25000 μg/ml

Now by using formula C₁V₁=C₂V₂ (Kanamycin),

C₁ = 25000 μg/ml,

V₁=?,

C₂= 100  μg/ml,

V₂=1000 ml

25000 μg/ml × V₁ = 100  μg/ml × 1000 ml

V₁ =  100  μg/ml × 1000 ml/25000 μg/ml

V₁ =   4 ml

Thus; in 1 lite of Lb+ Kan+Amp preparation;

0.5 ml of Amp & 4 ml of kanamycin is used for their stock preparation.

Finally;

Sterilization step should be carried out in flask (Clean dry glass wares) for media in an autoclave, the container size should be twice the volume of media which is prepared.

answerlinnaeus;