LAB 22: Genetic Transfer in Bacteria

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Questions and Answers

Which process involves bacteria taking in plasmids from their environment?

Binary Fission

Transformation (correct)

Transduction

Conjugation

What characterizes competent cells in bacterial transformation?

Cells that reproduce sexually

Cells that can take up plasmids (correct)

Cells capable of photosynthesis

Cells that are resistant to all antibiotics

What is the role of arabinose in the context of the pGLO gene?

It increases bacterial mutation rates

It induces the expression of the pGLO gene (correct)

It acts as a selective marker

It is necessary for conjugation

Which method would be used to make bacteria artificially competent?

Heat shock followed by ice treatment Signup and view all the answers

Which of the following describes transduction?

Transfer of DNA using a viral vector Signup and view all the answers

What type of gene does the pGLO plasmid confer resistance to?

Ampicillin resistance gene Signup and view all the answers

In which of the following plates would you expect to see growth of bacteria containing the pGLO plasmid?

LB +amp / +ARA Signup and view all the answers

What is the primary method of reproduction in bacteria?

Asexual reproduction through binary fission Signup and view all the answers

What is the outcome of conjugation in bacteria?

Exchange of plasmids between bacteria Signup and view all the answers

What occurs during the heat shock step of the transformation protocol?

The temperature rises to 42C for 50 seconds. Signup and view all the answers

Which of the following statements is true about horizontal gene transfer?

It allows for genetic recombination in bacteria Signup and view all the answers

What type of plates are used for the +pGLO tube after transformation?

LB + Ampicillin and LB + Ampicillin + Arabinose plates Signup and view all the answers

What is the purpose of adding LB to both tubes after the heat shock step?

To provide nutrients for bacterial recovery. Signup and view all the answers

What does the arabinose gene do in relation to the pGLO plasmid?

It allows GFP expression only in the presence of arabinose. Signup and view all the answers

What type of colonies are picked for the transformation process?

Single bacterial colonies from E. coli plates Signup and view all the answers

Which of the following statements accurately describes the -pGLO tube?

It is used to demonstrate the absence of transformation. Signup and view all the answers

Why is the heat shock procedure performed on the transformation solution?

To enhance the uptake of plasmid DNA by the bacteria. Signup and view all the answers

What role does the gene for ampicillin resistance play in the experiment?

It helps the bacteria survive in the presence of ampicillin. Signup and view all the answers

What is the significance of incubating the tubes on ice prior to heat shock?

It reduces bacterial metabolism to prepare for heat shock. Signup and view all the answers

What is the function of arabinose in the experiment described?

It serves as an inducer for the arabinose gene to express pGFP. Signup and view all the answers

What would be the predicted growth result of bacteria B on nutrient agar with vancomycin and streptomycin?

Growth Signup and view all the answers

Which media combination would NOT support the growth of bacteria B after conjugation with bacteria A?

Nutrient agar, +vanc, +strep, +merc Signup and view all the answers

What would be the expected result regarding pGFP expression in bacteria B after successful conjugation?

pGFP gene will express only in the presence of arabinose. Signup and view all the answers

What is the role of ampicillin in the media?

It serves as a selective agent against non-resistant bacteria. Signup and view all the answers

What type of bacteria is Species B before conjugation?

F- bacteria without a plasmid. Signup and view all the answers

If arabinose is present in the nutrient agar, what can be concluded about bacteria A?

Bacteria A will fluoresce regardless of other conditions. Signup and view all the answers

What does the term 'conjugation' refer to in the context of this experiment?

The transfer of genetic material between two bacteria. Signup and view all the answers

What role does the chromosomal gene for mercury resistance play in the experiment?

It has no impact on bacteria B. Signup and view all the answers

What would be the consequence of not adding arabinose to the media with bacteria A?

The pGFP gene will not be expressed. Signup and view all the answers

Study Notes

Transformation

Bacteria can acquire new genetic material through horizontal gene transfer
Horizontal gene transfer: the transfer of genetic information between adult organisms
Transformation: bacteria taking in plasmids from the environment
Transduction: transfer of DNA via bacteriophages
Conjugation: bacteria exchanging plasmids
Competent cells: bacteria that can be transformed by plasmids
Artificially competent cells: bacteria treated with heat shock to make them more permeable to plasmids
Heat shock process:
- Salt/heat to soften the cell membrane and allow plasmids to enter
- Ice to stiffen the membrane and trap the plasmids inside

pGLO Plasmid

pGLO plasmid contains two key genes:
- ampR gene: provides resistance to ampicillin
- pGLO gene: encodes the green fluorescent protein (GFP)
The pGLO gene is controlled by the pBAD promotor
pBAD promotor: requires arabinose to activate the pGLO gene
Arabinose: sugar molecule that acts as an inducer for the pGLO gene

Transformation Experiment

Aim: To transfer the pGLO plasmid containing the GFP gene into E. coli bacteria
Procedure:
- Two tubes labeled "-pGLO" and "+pGLO"
- "-pGLO" tube contained E. coli bacteria only
- "+pGLO" tube contained E. coli bacteria and the pGLO plasmid
- Both tubes were subjected to heat shock
- Each tube was plated on four different agar plates:
  - LB: nutrient-rich medium
  - LB + Amp: LB with ampicillin
  - LB + Amp + Ara: LB with ampicillin and arabinose
Expected results:
- -pGLO plates:
  - LB: bacterial growth
  - LB + Amp: no growth (ampicillin resistant genes absent)
- +pGLO plates:
  - LB + Amp: bacterial growth (ampicillin resistance due to plasmid)
  - LB + Amp + Ara: bacterial growth and green fluorescence under UV light (GFP expression induced by arabinose)

Conjugation Experiment

Aim: To transfer the pGLO plasmid from a donor bacterium (F+) to a recipient bacterium (F-)
Donor bacterium (F+):
- Contains the pGLO plasmid
- pGLO plasmid: ampicillin resistance, arabinose-inducible GFP, and GFP gene
- Contains a chromosomal gene for mercury resistance
Recipient bacterium (F-):
- Does not contain the pGLO plasmid
- Has chromosomal genes for streptomycin and vancomycin resistance
Expected results for recipient bacterium after conjugation:
- Nutrient agar, +vanc, +strep: growth (recipient bacterium retains its original resistance)
- Nutrient agar, +vanc, +strep, +amp: growth (recipient bacterium acquires ampicillin resistance from the plasmid)
- Nutrient agar, +vanc, +strep, +amp, +arab: growth and fluorescence (recipient bacterium acquires the pGLO plasmid and expresses GFP in the presence of arabinose)
- Nutrient agar, +vanc, +strep, +amp, +merc: no growth (recipient bacterium does not acquire mercury resistance as it is encoded on the donor's chromosome)

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Description

This quiz explores essential concepts of horizontal gene transfer in bacteria, with a focus on transformation, transduction, and conjugation. Additionally, it covers the properties of the pGLO plasmid and its components such as the ampR and pGLO genes. Test your understanding of these important microbiological processes.

LAB 22: Genetic Transfer in Bacteria - Transformation & pGLO