Lab Excersise 12 - MicroLab

Genetic Transformation and the UNiversal Genetic code

Define plasmid.

A small circular, double stranded DNA molecule that exists seperately from the bacterium’s main chromosome and replicates independently each time the cell divides passing the foreign gene to every daughter cell.

The plasmid is passed to EVERY daughter cell.

True

What is the Origin of Replication (Ori)

the sequence of DNA where replication begins, allowing the plasmid to copy itself inside the bacterial cell independently of the chromosome.

What is the Antibiotic Resistance Gene (bla-amp^R)?

codes for beta-lactamase. Is always expressed that allows transformed bacteria to survive on ampicillin-containing media.

What enzyme does the bla-ampR gene code for?

Beta-lactamase

What is the gene of interest (GFP)?

codes for green flourescence protein and will only be produced when the promoter is active.

What does GFP stand for?

green flourescnce protein

What is the inducible promoter (P*)?

controls the expression of the GFP gene.

What switches on the inducible promoter P*?

arabinose (sugar/carb)

What would happen if arabinose is not present?

GFP is not produced.

What would happen if arabinose is present?

the promoter is activated and the bacteria expresses GFP, causing them to glow green in UV light.

Bacteria normally take up DNA from their environment.

False

Why can DNA not pass into the bacterial cell normally?

the cell membrane acts as a barrier and the DNA is too negatively charged.

Define a competent cell.

a bacteria that has been treated to make it’s membrane more permable.

What does calcium chloride do for E.coli?

nuetralizes the negative charge on both DNA and the cell surface, reducing repulsion and allowing DNA to associate with the outer membrane when given the right conditions.

What does heat shock do for the bacteria?

opens up the membrane allowing the DNA to pass in, but the heat nearly kills the bacteria. Putting it back in the ice water constricts the membrane now with the DNA inside.

Every cell takes in the plasmid.

False

If you do not see growth on the plate, what does this imply?

the bacteria did not transform and were killed by the antibiotic