Download Antibiotics - West Chester University of Pennsylvania

Antibiotics
Ampicillin and Kanamycin
Ampicillin
-Inihibits cell wall synthesis
-Irreversible inhibits transpeptidase, which cross-links
the peptidoglycan
Ampicillin Resistance
-beta-lactam antibiotic
-Acquires a gene that encodes β-lactamase
-Cleaves β-lactam ring of
ampicillin
-Ampicillin is no longer
bound to transpeptidase
and therefore functions
normally
Kanamycin
-an aminoglycoside (amino modified sugar) bacteriocidal
antibiotic
-Isolated from Streptomyces kanamyceticus
-Binds to the 30S procaryotic
ribosome
Kanamycin
-Binds to the 30S procaryotic
ribosome
Kanamycin Resistance
-aminoglycoside modifying enzymes, phosphorylation,
making Kanamycin uneffective
Bacteriostat vs. Bacteriocide
Ampicillin – inhibits new cell wall synthesis
Kanamycin – inhibits protein synthesis
Restriction Endonucleases
Nuclease – Enzymes that digest or cut DNA and RNA
Endonuclease vs. Exonuclease
Endonuclease – Cut nucleic acids at sites internal to
the molecule
Exonuclease - Cut nucleic acids at sites external to the
molecule
DNA Properties
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Restriction Endonuclease
-1950s, a “primitive immune system” in E. coli in
defense of bacteriophage lambda infection
-1968, Restriction enzyme HindIII was isolated from
Haemophilus influenzae
-Recognition site for HindIII
5’ G T A C 3’
3’ C A T G 5’
Restriction Endonuclease
-1978, Nobel prize awards for the discovery of
restriction endonucleases and their application to
molecular genetics
-Currently over 3000 restriction endonulceases
isolated with 230 recognition sequences
-Mostly isolated from bacteria, but some from
viruses, archaea, and eucaryotes
HindII
-Recognition site for HindIII
5’ A A G C T T 3’
3’ T T C G A A 5’
-Cleavage Products
5’ A
3’ T T C G A
A G C T T 3’
A 5’
Lambda (λ) Phage
-Bacteriophage with double-stranded DNA genome, 48,502 bp
DNA Gel Electrophoresis
-Separation of DNA
fragments through agarose
by application of an electric
field
DNA Gel Electrophoresis
DNA Gel Electrophoresis
-DNA Mobility is
inversely proportional to
Log10 of its Molecular
Weight