Name: Lilli Crawford HOMEWORK: LB-agar plate preparation (4 points)
Section: M/W PM
Directions: This assignment is worth 4 points. Use whatever resources are available on eLC but paraphrase
what you find by responding in your own words.
1. Regarding LB-agar plates
a. What role does an LB-agar plate perform in working with bacterial cultures?
LB-agar plates allow us to isolate desired individual colonies of bacteria.
b. What does LB stand for? What role does it play in working with bacterial cultures?
Lysogenic broth; LB is used as a source of food for bacteria and allows us to grow a large
amount of a specific bacteria to study.
2. Why is it that when a scientist needs only a few nutrient agar plates, they prepare many more than
this?
Scientists prepare multiple agar plates at once because it is handy to have extra on hand and the
sterilization process can cause evaporation. If you are preparing a small volume of LB-agar, a
large portion will evaporate, leaving you with not enough to make a plate.
3. A student needs to do a “protein prep”, which means that the student needs to express and purify a
recombinant protein from E. coli. The student has the gene of interest inside an expression
plasmid vector, which is located inside E. coli that is stored in a -80oC freezer. The student
thawed a vial of E. coli and streak the cells onto an LB-agar plate containing an antibiotic.
a. What is the function of the antibiotic?
The antibiotic allows for selection of only the bacteria with resistance to the
antibiotic.
b. The antibiotic is added to the molten LB-agar solution before it solidifies. However, the
antibiotic is temperature sensitive and becomes inactivated if the temperature exceeds
50-55oC. temperature sensitive. Use the table below to describe the results you expect to
see, in terms of bacterial growth bacterial growth, if the antibiotic was added when the
temperature was much greater than 50oC or roughly equivalent to 50-55oC.
Expected results if added at a temperature
between 50-55oC
Expected results if added at temperature
much greater than 50-55oC
We will see antibiotic selection and only
the colonies which are resistant will
survive.
There will be no antibiotic selection due
to the denaturing of the antibiotic. All
colonies will survive.
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