UNIT 9: MICROBIAL GENETICS & BIOTECHNOLOGY

LEARNING OBJECTIVES
Upon successful completion of this unit, you will be able to:
1. define the following terms: genetic control, operon, operator, promoter, and regulator gene.
2. compare and contrast genetic control enzyme systems using the lac and trp operons as examples.
3. summarize the three main mechanisms of DNA transfer in bacteria.
4. define and identify the following terms and concepts: recombination, prototroph, auxotroph, donor, recipient, genetic markers,
wildtype, selection and counter selection.
5. interpret results of a genetic mapping experiment including identifying appropriate donor, recipient, and recombinant strains
along with appropriate media components.
6. explain how transformation and genetic control are used in genetic engineering.
7. define the following terms: biotechnology, genetic engineering, genetically modified organisms (GMOs), sticky ends,
restriction site, selectable marker and screenable marker.
8. outline the steps in making recombinant DNA and genetically modified organisms. Provide examples uses of genetically
modified bacteria, viruses, plants, and animals.
9. describe the roles of PCR, gel electrophoresis, restriction enzymes, plasmid, DNA ligase, selective and differential media
in genetic engineering.
10. explain how microarray chip analysis are used to identify pathogens in patient samples.
11. briefly describe how CRISPR works and the future applications of this new technology.

REFERENCES
Chapter 11 and 12

SAMPLE PROBLEMS
1. Can two observably different cells have the same genes? Explain. (9-1; A)

2. You are working with a genetic mapping experiment by conjugation of a newly discovered microorganism. You already know the
organism has the following genes for biomolecule production: lys (lysine), leu (leucine), ilv (isoleucine), tyr (tyrosine), ser (serine)
and gly (glycine). Also you have chloramphenicol and tetracycline antibiotic resistant strains of this organism (CmR and TetR). To
keep your experiment simple, you must choose 4 biomolecule production genes and 1 antibiotic to study. (There are multiple
answers to this problem. I am looking for you to synthesize your own experiment.) (9-4, 5; E)
a. Describe the genetic make up of the donor strain you’ve chosen to use in your experiment.

b. Describe the genetic make up of the recipient strain you’ve chosen to use in your experiment.

c. Describe the best media for selecting only recombinant cells.

d. Fill in the below table with media components along the top row. In each of these cases, the minimal media has ALL
amino acids except the specific one you’ve listed next to the minus sign.
Time

0 NG NG NG NG
2 NG G NG NG
4 NG G NG NG
6 NG G NG G
8 NG G G G
10 G G G G

e. Using your filled in table, what is the order of the genes used in your experiment?
 3. Is biotechnology always associated with genetic engineering? Explain your answer. (9-7; U)

4. What are some advantages of cloning human genes into bacteria to treat human diseases caused by specific protein deficiencies?
(9-8; U)

(9-5; A) 1. Some bacteria undergo recombination by a process called conjugation. During conjugation, a portion of the DNA from
the donor cell is transported into a recipient cell. Suppose you have donor cells, which are prototrophs for leucine
synthesis (the gene is represented as leu +) and are sensitive to the antibiotic streptomycin. What would be the most
appropriate recipient cells for conjugation?
a. Leucine prototrophs (leu +) that are resistant to streptomycin.
b. Leucine prototrophs (leu +) that are sensitive to streptomycin.
c. Leucine auxotrophs (leu -) that are resistant to streptomycin.
d. Histidine auxotrophs (his -) that are resistant to streptomycin.
e. Histidine prototrophs (his +) that are resistant to streptomycin.

(9-5; A) 2. After you have mixed the two kinds of bacteria from Question #1 and allowed them to conjugate, what kind of medium
would you use to identify only those bacteria that have undergone genetic recombination?
a. A complete medium with streptomycin.
b. A complete medium without streptomycin.
c. An incomplete medium (lacking leucine) with streptomycin.
d. An incomplete medium (lacking histidine) with streptomycin.
e. An incomplete medium (lacking both histidine and leucine) with streptomycin.

(9-5; E) 3. You set up an experiment to study conjugation between two strains of bacteria. The donor strain is prototrophic for
the chromosomal genes A+, B+, C+, and D+. The recipient strain is auxotrophic for the same genes (A-, B-, C-, and D-).
You interrupt the conjugation process for a portion of the cells at two minute intervals, and grow these on appropriate
media to detect recombination for each of the genes. The results are plotted below. In what order are the genes found
on the bacterial chromosome?
a. A, B, C, D
b. D, C, A, B
c. B, C, A, D
d. C, B, D, A
e. B, A, D, C

(9-9; A) 4. The restriction enzyme TaqI cleaves the nucleotide sequence TCGA between the T and C on both strands. Assume you
have cut vector DNA with TaqI. Which represents an end(s) of the DNA to be cloned that will base pair with the “sticky
ends” produced by TaqI cleavage?
a. T c. CGA
AGC T
b. TCG d. “a” and “b” will base pair with the “sticky ends”
A e. “a” and “c” will base pair with the “sticky ends”
(9-9; U) 5. In a recombinant DNA experiment, what would be the consequence of the restriction site being located in the
selectable marker (an antibiotic resistance gene)?
a. Only cells receiving the plasmid with the gene of interest will grow on the selective medium.
b. Only cells receiving the plasmid without the gene of interest will grow on the selective medium.
c. Only cells not receiving the plasmid will grow on the selective medium.
d. All cells receiving the plasmid–with or without the gene of interest–will grow on the selective medium.

(9-9; U) 6. PCR is a technique that replaces

a. electrophoresis. d. autoradiography.
b. cDNA production. e. gene therapy.
c. DNA plasmid cloning.

(9-8; A) 7. Assume you have at your disposal a plasmid to be used as a vector in a recombination experiment in which E. coli is
the recipient. The plasmid already has the gene to be cloned inserted into it. It also has a streptomycin resistance
gene, a lac regulatory gene, a lac promoter and a lac operator in it. The lac genes are located next to the gene to be
cloned. Which strain of E. coli should be used as the recipient in this experiment?
a. An E. coli that is resistant to streptomycin.
b. An E. coli that is sensitive to streptomycin.

(9-8; A) 8. Which medium should be used to isolate recombinant cells in the experiment outlined in Question #8?
a. nutrient agar plus lactose
b. nutrient agar without lactose and without streptomycin
c. nutrient agar without streptomycin; lactose is irrelevant
d. nutrient agar with streptomycin; lactose is irrelevant

(9-8; A) 9. Once you have isolated the recombinant cells in Question #8, you transfer them to a broth to grow a high cell density.
Then, when you are ready for them to start production of the cloned gene product, what should you add to the
medium?
a. lactose
b. streptomycin
c. lactose and streptomycin

(9-2; U) 10. Which explains why transcription is turned OFF when lactose is present?
a. The repressor binds to the operator blocking DNA polymerase.
b. The repressor binds to the operator blocking RNA polymerase.
c. The repressor binds to the RNA polymerase inactivating it.

(9-4; U) 11. What do F+ cells have that other cells do not have?
a. The F plasmid
b. The conjugative pilus
c. A lysogenic phage
d. Options A & B
e. Options B & C

(9-4; U) 12. What will grow on Minimal media without threonine and with streptomycin (MM –Thr + Strep)?
a. Donor Hfr cell
b. Recipient cell
c. Recombinant cell
d. All of these will grow.

(9-10; U) 13. How does Cas9 know where to cut?

a. The guide RNA base pairs to the targeted DNA.
b. By cutting the DNA first.
c. The enzyme works randomly cutting all 3 nucleotide locations.
d. CRSIPR looks for the proteins involved in mutation causation.