Microbiology Lecture 6 Overview Chapter 6; Microbial Genetics

Griffith and Avery, McCarty, and MacLeod experiments

Overview of DNA structure

G+C content: Centrifugation; melting temperature to determine.

Similar G+C not necessarily related

Semiconservative replication; Messelson/Stall experiment

Overview of DNA simultaneous, bidirectional replication. Leading/lagging strands

Transcription

Translation

Regulation; constitutive, inducible, repressible

"Tell me about the Lac Operon!"

Attenuation

Global regulation; CAP

Plasmids; F., Hfr., R., catabolic, toxin-encoding.

Mutations; missense, nonsense, frame-shift, addition, deletion, +p.171

Mutagens and carcinogens; Ames' Test

Mutagenesis and selection (direct, counterselection, brute force, site-directed)

Replica plating; toothpick technique

Recombination; transformation, conjugation, transduction (specialized, generalized).

Lytic and lysogenic cycles; cotransduction, cotransformation, interrupted mating

Mechanisms of antibiotic resistance

New goal: sequence your genome for $1,000! (Billions for first sequence)

http://web.mit.edu/esgbio/www/pge/lac.html

Above obtained as one of 10,000 hits in search for lac operon

The Lac Operon

Jacob and Monod were the first scientists to elucidate a transcriptionally regulated system. They worked on the lactose metabolism system in E. Coli. When the bacterium is in an environment that contains lactose: It should turn on the enzymes that are required for lactose degradation. These enzymes are:

beta-galactosidase:

This enzyme hydrolyzes the bond between the two sugars, glucose and galactose. It is coded for by the gene LacZ.

Lactose Permease:

This enzyme spans the cell membrane and brings lactose into the cell from the outside environment. The membrane is otherwise essentially impermeable to lactose. It is coded for by the gene LacY.

Thiogalactoside transacetylase:

The function of this enzyme is not known. It is coded for by the gene LacA.

These three enzymes appear adjacent to each other on the E. Coli genome. They are preceded by a region which is responsible for the regulation of the lactose metabolic genes. Note that there is more to regulation than the obvious. It would seem that the cell would want to turn these genes on when there is lactose around and off when lactose is absent. But the story is more complicated than that.

A bacterium's prime source of food is glucose, since it does not have to be modified to enter the repiratory pathway. So if both glucose and lactose are around, the bacterium wants to turn off lactose metabolism in favour of glucose metabolism. There are sites upstream of the Lac genes that respond to glucose concentration. This assortment of genes and their regulatory regions is called the Lac operon. The Lac operon has been examined in detail. Here is a diagram of it: