The Escherichia Coli Genome

Lecture Exam 2

Escherichia Coli (E. Coli)

gram negative bacillus shaped bacterium that is commonly found in the lower intestines of warm-blooded organisms

- 4.64 Mbp

- 4300 genes

how many Mbp is the E. Coli genome? how many genes?

plasmid

small circular double stranded DNA molecules that replicate independently of the host chromosome; provides antibiotic resistance

conjugation

how are plasmids often transferred between bacteria?

tra gene - builds the sex pilus (thin protein tube connecting the 2 bacteria) + contains other necessary materials

what gene controls the transfer of plasmids? how?

-contains genes that encode resistance to antibiotics

-easily transfer antibiotic resistance to other bacteria

what are 2 characteristics of R-plasmids?

-27F (binds at beginning)

-1492R (binds at the end)

both are used to amplify the 16S gene

what primers are typically used in PCR? why are these primers used?

it is present in all bacteria with regions specific to different bacteria, making the type more identifiable

what is the benefit of amplifying the 16S gene?

1. denaturation (94-96°C) 2 min

2. annealing (40-70°C) 30 sec

3. elongation (72°C) 1.5 min

repeated 34 times

what are the 3 steps to PCR (+ temp. and time)? how many times is this process repeated?

-DNA template

-dNTP

-polymerase

-buffer

-primers

-water

-Taq polymerase

what is required for PCR? (7)

Kary Mullis

applications:

-phylogenetic studies

-identification of microbes

-forensics

who invented PCR? what are its 3 applications?

agarose gel electrophoresis

technique used to visualize and separate DNA and PCR products

an electric current moves charged DNA molecules through the gel matrix; small m-c move faster (will appear farther down)

how does agarose gel electrophoresis work?

agarose

polysaccharide extracted from seaweed that dissolves in water and forms a gel when cooled