Bacterial Conjugation and Transposon Mutagenesis Notes

A set of vocabulary flashcards derived from lecture notes on bacterial conjugation, transposon mutagenesis using plasmid pRL27, and the phenotypic screening of S. marcescens variants.

$$pRL27$$

The specific plasmid used in the experiment, originally found in $E.\,coli\,BW20767$.

Transposase

An enzyme that recognizes inverted repeats ($IRs$) and catalyzes the movement or insertion of a transposable element.

Inverted repeats ($IRs$)

The specific DNA sequences that transposase recognizes and binds to for excision and insertion.

$$TetAp$$

The component of the $pRL27$ plasmid that functions as a promoter.

$$tnp$$

The gene on the $pRL27$ plasmid that codes for transposase.

$$aph$$

The gene on the $pRL27$ plasmid responsible for kanamycin resistance.

$$oriT$$

The origin of transfer sequence located on the $pRL27$ plasmid.

$$oriR6K$$

The origin of replication found within the $pRL27$ plasmid.

Conjugation

The process of transferring DNA between bacteria through direct cell-to-cell contact.

$$E.\,coli\,BW20767/pRL27$$

The donor bacterium used in the conjugation experiment.

$$S.\,marcescens$$

The recipient bacterium intended to receive the $pRL27$ plasmid to disrupt its prodigiosin pathway.

Kanamycin

An antibiotic used to select for bacteria that successfully carry the $pRL27$ plasmid.

$$MOPS$$

A minimal medium used to select against the $E.\,coli\,BW20767/pRL27$ donor strain.

Auxotroph

An organism, such as the $E.\,coli$ donor strain, that requires a specific nutrient (like leucine) that the wild-type strain does not.

Leucine

The specific amino acid missing from $MOPS$ medium, preventing the growth of the $E.\,coli$ leucine auxotroph.

Serial dilutions

A technique performed using $10^{-3}$, $10^{-4}$, and $10^{-5}$ concentrations to obtain isolated colonies for phenotypic analysis.

Prodigiosin

The red pigment produced by $S.\,marcescens$, used as a marker for phenotypic screening.

White colony phenotype

Indicates a complete disruption of the prodigiosin pathway, often occurring if the transposon inserts early in the sequence.

Dark pink colony phenotype

Indicates less disruption of the prodigiosin production, suggesting the pathway was disrupted late.

Phenotypic screening

The process of identifying and comparing mutant phenotypes (dark pink, light pink, and white) with the wild-type strain.