$$\text{DNA (GFP gene)} \xrightarrow{\text{RNA polymerase}} \text{mRNA} \xrightarrow{\text{ribosome}} \text{GFP protein}$$

Question 1

Which gene on the pGLO plasmid is responsible for making bacteria resistant to ampicillin, and how does it confer this resistance?

Accepted Answer

The bla gene; it encodes beta-lactamase, which breaks down ampicillin.

Question 2

In the pGLO transformation experiment, what is the role of arabinose in the growth medium?

Accepted Answer

It binds to the araC protein, activating transcription of the GFP gene.

Question 3

Why do only a small percentage of bacteria become transformed during the heat shock step?

Accepted Answer

Only a few bacteria take up the plasmid DNA due to temporary membrane permeability.

Question 4

If you plate transformed bacteria (+pGLO) on LB/amp/ara plates, what phenotype do you expect to observe under UV light?

Accepted Answer

Colonies will grow and fluoresce green.

Question 5

What would you expect to see if you plated non-transformed bacteria (-pGLO) on LB/amp plates?

Accepted Answer

No bacterial growth.

Question 6

Which of the following best describes the function of the ori region on the pGLO plasmid?

Accepted Answer

It is the origin of replication, allowing the plasmid to replicate independently.

Question 7

Why is it important to include a (-)pGLO control in the transformation experiment?

Accepted Answer

To demonstrate that only bacteria with the plasmid can survive on ampicillin plates.

Question 8

Which step in the transformation protocol is critical for making the bacterial cell membrane permeable to plasmid DNA?

Accepted Answer

Heat shock at 42°C.

Question 9

If you observe white colonies on an LB/amp plate from the (+)pGLO transformation, what is the most likely explanation?

Accepted Answer

The GFP gene is not being expressed due to the absence of arabinose.

Question 10

Which of the following best explains why the bla gene is described as being 'constitutively expressed'?

Accepted Answer

It is always expressed, regardless of environmental conditions.

Question 11

A student forgets to add arabinose to the LB/amp plate when plating (+)pGLO bacteria. What will be the result under UV light?

Accepted Answer

Colonies will grow but will not fluoresce.

Question 12

Which of the following equations best represents the process of transcription required for GFP expression in the presence of arabinose?

Accepted Answer

$$	ext{DNA (GFP gene)} \xrightarrow{	ext{RNA polymerase}} 	ext{mRNA} \xrightarrow{	ext{ribosome}} 	ext{GFP protein}$$

Question 13

Why do scientists use the GFP gene as a marker in transformation experiments?

Accepted Answer

It allows for easy visual identification of transformed cells under UV light.

Question 14

If you observe growth on the (-)pGLO LB plate but not on the (-)pGLO LB/amp plate, what does this indicate?

Accepted Answer

The bacteria require the pGLO plasmid to survive in the presence of ampicillin.

Question 15

Which of the following best describes the regulatory relationship between araC, arabinose, and GFP expression in the pGLO system?

Accepted Answer

araC activates GFP expression only when bound to arabinose.

Question 16

A researcher wants to select only bacteria that have taken up the pGLO plasmid. Which plate should they use?

Accepted Answer

LB/amp plate.

Question 17

Which of the following steps is NOT necessary for the successful transformation of bacteria with pGLO?

Accepted Answer

Adding arabinose to the transformation solution before heat shock.

Question 18

If a mutation occurred in the araC gene on the pGLO plasmid, which outcome would you most likely observe?

Accepted Answer

Bacteria would not fluoresce green, even in the presence of arabinose.

Question 19

Which of the following best explains why only transformed bacteria grow on LB/amp plates?

Accepted Answer

Only transformed bacteria express beta-lactamase, which inactivates ampicillin.

Bacterial Transformation and the pGLO Plasmid: Principles and Laboratory Procedure

Study Guide - Practice Questions