MICRO Module 5 Flashcards
(30 cards)
What is transformation in bacteria?
Transformation is the process by which bacteria acquire new genetic material (DNA) from their environment, allowing them to adapt to environmental changes.
What is horizontal gene transfer?
Horizontal gene transfer refers to the movement of DNA between bacteria that are not parent and offspring, as seen in transformation.
What are competent bacteria?
Competent bacteria are those that can readily take up naked DNA from their environment.
What are plasmids?
Plasmids are circular, double-stranded DNA molecules used in transformation to carry genes of interest.
How do plasmids help in transformation?
Plasmids can be easily manipulated to carry genes and can replicate independently within bacterial cells.
What is the purpose of antibiotic resistance genes in plasmids?
Antibiotic resistance genes allow transformed bacterial cells to survive in the presence of antibiotics.
What is the role of the pGFP plasmid in the experiment?
The pGFP plasmid introduces two traits into E. coli: green fluorescence from the GFP gene and resistance to Ampicillin.
What is the significance of heat shock in transformation?
Heat shock generates thermal currents that facilitate the uptake of DNA through pores in the bacterial cell membrane.
What is the expected outcome of the transformation experiment?
The expected outcome is the growth of colonies on selective media containing Ampicillin, indicating successful transformation.
Why are controls important in an experiment?
Controls are essential to validate the results and ensure that the observed effects are due to the experimental treatment.
What should be done to isolate transformed cells?
Transformed cells can be isolated by growing them on media containing antibiotics, allowing only those that have taken up the plasmid to survive.
What is the incubation temperature for E. coli after adding nutrient broth?
The incubation temperature for E. coli after adding nutrient broth is 37°C for 30 minutes.
What should be observed on the plates after incubation?
Colony growth should be observed on the plates, with differences in colony color and number based on DNA uptake.
What is the purpose of plating cells without pGFP DNA?
Plating cells without pGFP DNA serves as a control to assess the baseline growth of E. coli without transformation.
How can you differentiate between transformed and non-transformed cells?
Transformed cells will express the GFP gene, resulting in green fluorescence under UV light, while non-transformed cells will not.
What is the expected colony count on plates without Ampicillin?
Plates without Ampicillin should show a high number of colonies, as there is no selection pressure against non-transformed cells.
What plasmid was used to transform E. coli?
The plasmid used was pGFP (0.05 µg).
What was the purpose of the heat shock in the transformation protocol?
The heat shock was applied to facilitate the uptake of the plasmid DNA by E. coli.
What was the final volume of broth used to resuspend the cells after heat shock?
The final volume of broth used was 0.5 mL.
How much of the resuspended cells was plated on nutrient agar?
0.25 mL of the resuspended cells was plated on nutrient agar.
What were the two types of nutrient agar plates used in the experiment?
The two types of plates were those containing Ampicillin (AMP, 100 microgram/mL) and those without Ampicillin.
What was the expected result for the NA without AMP plate?
Expected results: 10,000 white colonies under visible light and 0 green colonies under UV light.
What was the expected result for the NA with AMP plate without pGFP?
Expected results: 0 white colonies and 0 green colonies.
What was the expected result for the NA without AMP plate with pGFP?
Expected results: 10,000 white colonies and 100 green colonies.
