Genetics - Lecutre 11: Bacterial Genetics and Jacob and Monod and the lac Operon

What do bacterial colonies lack?

differences in phenotypes

Prototroph

a microorganism that does not require any organic growth factors (E.coli)

Auxotroph

cannot survive on minimal media

Benefit of replica plating

allows us to copy one set of colonies onto multiple different media and determine mutations therein

Bernard Davis experiment

Physical contact between the two strains was needed for recombinant cells to form.

Pilli

Hollow tubes used to move cells or exchange DNA between bacteria by conjunction.

Bacterial gene transfer is...

unidirectional

Fertility factor (F factor)

A specific plasmid (transferred by an F+ donor cell) that contains the genes needed for pilus formation and DNA export.

Hfr strains

undergo recombination at high frequency, but recipient cells generally do not become donors

Why can't recipients of Hfr strains become donors?

Hfr strains have the F-factor integrated onto their main chromosome

Why does an F-factor integrated onto the main genome prevent recipient cells from becoming donors?

There is no plasmid to be transferred through the pillus, so the entire chromosome is sent through, but the pillus is fragile and often breaks during the transfer, leaving an incomplete, linear piece of DNA in the recipient.

Exogenote

linear piece of chromosome transferred into a bacterial cell from another cell by conjugation

What happens to an exogenote?

If genes on the exogenote are to persist, they need to be recombined into the existing, circular chromosome. This is done by double recombination.

Fragility of pillus and mapping genes:

crossed certain strains of bacteria that were resistant to different antibiotics and found it took around 2 hrs for entire genome to transfer, and for daughter cells to be resistant to all three antibiotics. Then transfer was arrested in different stages using a blender to observe results

Based on pillus mapping data, what conclusion was drawn about the shape of the bacterial chromosome?

It was circular

Why does an exogenote need to undergo double recombination to be viable?

Without double recombination, the chromosome would become linear, which is not viable in bacteria

Messy excision of integrated Hfr:

F' plasmid can have other genes from the chromosome that are not usually donatable

Episome

plasmid that can integrate into bacterial chromosome

Merodiploid

a bacterial cell that is diploid for a portion of its genome

Why is the lac operon a good model system?

beta-gal is ONLY made in the presence of lactose - postive and negative regulation

How can beta-gal activity be monitored?

Processing known substrates into colorful derivatives that act as indicators

Inducers

Compounds that stimulate the transcription of specific genes

IPTG

inducer of lac operon: inactivates repressor but is not broken down by β-galactosidase

lacZ

encodes B-galactosidase

lacY

encodes lactose permease

Describe the relationship between lacZ and lacY

co-regulated

lacI

encodes the lac repressor

lacI-

mutation that causes cells to produce enzymes all the time (constitutively)

lacI^S mutation

super repressor. cannot bind lactose or IPTG and therefore cannot leave operator so the system is always shut off.

lacO

operator where the repressor binds

2 classes of lacO mutations:

sometimes they were constitutive (O^c)

sometimes they genes following were permanently turned off

lacP

lac promoter

Mutation in lacP (lacP-)

genes will never be expressed

Structural genes of lac operon

lacZ, lacY, lacA

Which part of the lac operon is cis-dominant?

lacO

Which part of the lac operon is trans-dominant?

lacI