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Game Plan

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Biofilm - organized microbial system of layers of microbial cells ... to sessile (i.e. what causes biofilm formation)? Genes being turned on and off! Example: ... – PowerPoint PPT presentation

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Title: Game Plan


1
Game Plan
Lab Review temp and UV results Chemical
growth control Streak plates Prelab Open Lab
Lecture Biofilms Review of basic
genetics Bacterial gene structure Gene
regulation Mutations
2
Biofilms
  • Biofilm - organized microbial system of layers of
    microbial cells embedded in a polysaccharide
    matrix of microbial origin associated with
    surfaces
  • Medicine - on catheters, teeth, middle ear
    infections, GI, GU tract, lungs (cystic
    fibrosis), biomedical products
  • Aquatic environments - on algae, rocks, ships
  • Industry - pipes, air conditioning vents, plastics

3
Biofilms
Industrial biofilm
Dental biofilm
Intestinal biofilm
4
Biofilm formation
5
Why form a biofilm?
1. Protection from antibiotics, toxins, and
immune cells
Effect of tobramycin (A) and ciprofloxacin (B) on
survivial of planktonic cells
(circle) resuspended biofilm cells
(triangle) biofilm colony cells (squares) Open
symbols are untreated
6
Why form a biofilm?
1. Protection from antibiotics, toxins, and
immune cells 2. Source of nutrients/ limits on
nutrients
7
Why form a biofilm?
  • 1. Protection from antibiotics, toxins,
  • and immune cells
  • 2. Source of nutrients/ limits
  • on nutrients
  • 3. Favorable microenvironment
  • Highly hydrated
  • Low oxygen

8
Why form a biofilm?
  • 1. Protection from antibiotics, toxins,
  • and immune cells
  • 2. Source of nutrients/ limits
  • on nutrients
  • 3. Favorable microenvironment
  • Highly hydrated
  • Low oxygen
  • 4. Stability - can detach and leave

Figure 8.1a
9
Why form a biofilm?
  • 1. Protection from antibiotics, toxins,
  • and immune cells
  • 2. Source of nutrients/limits
  • on nutrients
  • 3. Favorable microenvironment
  • Highly hydrated
  • Low oxygen
  • 4. Stability - can detach and leave
  • 5. Community
  • Gene transfer, signal transduction, quorum sensing

10
Review of DNA processing
Figure 8.2
11
Genetics terminology
Genetics- the study of genes Genes- a segment of
DNA that codes a functional production
(protein) Genome- all of the genetic material
in a cell Genomics- molecular study of
genomes Genotype- genes of an organism
Phenotype- physical expression of the genes
12
DNA structure
Figure 8.3b
13
DNA replication
  • Enzymes DNA polymerase, DNA ligase
  • primase to make RNA primers,
  • accessory enzymes (topoisomerase,
  • gyrase, helicase)
  • Antiparallel 5 to 3 synthesis results in
  • leading and lagging strands
  • Semi-conservative replication

Figure 8.5
14
Bacterial DNA replication
Figure 8.6 - Overview
15
Pit stop
The E. coli genome replicates every 45 minutes,
but divides every 26 minutes in ideal
conditions. How do you reconcile these two
facts?
16
RNA transcription
  • Enzymes RNA polymerase
  • Promoters and terminators start and end
  • process
  • 5 to 3 synthesis
  • Types tRNA, mRNA, rRNA

Figure 8.7
17
RNA processing in eukaryotes
18
Translation and the Genetic Code
  • Players ribosomes ( rRNA), tRNA,
  • and mRNA
  • Codons
  • Anticodons

19
Translation and the Genetic Code
  • Players ribosomes ( rRNA), tRNA,
  • and mRNA
  • Codons
  • Anticodons

20
Translation
21
Translation
22
Universal Genetic Code
Gene 5 ATG GTC CGA GCC CGC TAA GGC 3
3 TAG CAG GCT CGG GCG ATT CCG 5
Figure 8.8
23
Universal Genetic Code
Gene 5 ATG GTC CGA GCC CGC TAA GGC 3
3 TAG CAG GCT CGG GCG ATT CCG 5 RNA 5 AUG
Figure 8.8
24
Universal Genetic Code
Gene 5 ATG GTC CGA GCC CGC TAA GGC 3
3 TAG CAG GCT CGG GCG ATT CCG 5 RNA 5 AUG GUC
Figure 8.8
25
Universal Genetic Code
Gene 5 ATG GTC CGA GCC CGC TAA GGC 3
3 TAG CAG GCT CGG GCG ATT CCG 5 RNA 5 AUG
GUC CGA
Figure 8.8
26
Universal Genetic Code
Gene 5 ATG GTC CGA GCC CGC TAA GGC 3
3 TAG CAG GCT CGG GCG ATT CCG 5 RNA 5 AUG
GUC CGA GCC
Figure 8.8
27
Universal Genetic Code
Gene 5 ATG GTC CGA GCC CGC TAA GGC 3
3 TAG CAG GCT CGG GCG ATT CCG 5 RNA 5 AUG
GUC CGA GCC CGC
Figure 8.8
28
Universal Genetic Code
Gene 5 ATG GTC CGA GCC CGC TAA GGC 3
3 TAG CAG GCT CGG GCG ATT CCG 5 RNA 5 AUG
GUC CGA GCC CGC UAA
Figure 8.8
29
Universal Genetic Code
Gene 5 ATG GTC CGA GCC CGC TAA GGC 3
3 TAG CAG GCT CGG GCG ATT CCG 5 RNA 5 AUG
GUC CGA GCC CGC UAA GGC 3 Protein

Figure 8.8
30
Universal Genetic Code
Gene 5 ATG GTC CGA GCC CGC TAA GGC 3
3 TAG CAG GCT CGG GCG ATT CCG 5 RNA 5 AUG
GUC CGA GCC CGC UAA GGC 3 Protein
Met
Figure 8.8
31
Universal Genetic Code
Gene 5 ATG GTC CGA GCC CGC TAA GGC 3
3 TAG CAG GCT CGG GCG ATT CCG 5 RNA 5 AUG
GUC CGA GCC CGC UAA GGC 3 Protein
Met Val
Figure 8.8
32
Universal Genetic Code
Gene 5 ATG GTC CGA GCC CGC TAA GGC 3
3 TAG CAG GCT CGG GCG ATT CCG 5 RNA 5 AUG
GUC CGA GCC CGC UAA GGC 3 Protein
Met Val Arg
Figure 8.8
33
Universal Genetic Code
Gene 5 ATG GTC CGA GCC CGC TAA GGC 3
3 TAG CAG GCT CGG GCG ATT CCG 5 RNA 5 AUG
GUC CGA GCC CGC UAA GGC 3 Protein
Met Val Arg Ala
Figure 8.8
34
Universal Genetic Code
Gene 5 ATG GTC CGA GCC CGC TAA GGC 3
3 TAG CAG GCT CGG GCG ATT CCG 5 RNA 5 AUG
GUC CGA GCC CGC UAA GGC 3 Protein
Met Val Arg Ala Arg
Figure 8.8
35
Universal Genetic Code
Gene 5 ATG GTC CGA GCC CGA TAA GGC 3
3 TAG CAG GCT CGG GCT ATT CCG 5 RNA 5 AUG
GUC CGA GCC CGA UAA GGC 3 Protein
Met Val Arg Ala Arg Stop (formylmethionin
e in bacteria)
Figure 8.8
36
DNA processing in bacteria- simultaneous
transcription/translation
37
Game Plan
Lecture Gene regulation and the
operon Mutations Bring books next class
for APO-2
Lab Open Lab LAB EXAM NEXT CLASS
38
Review of DNA processing
Figure 8.2
39
What changes a bacterium from planktonic to
sessile (i.e. what causes biofilm formation)?
Genes being turned on and off! Example Different
ial expression of 2.9-17 P. aeruginosa genes
between planktonic and biofilm cells
40
When are genes on and off?
  • Constitutive genes genes are constantly on
    (60-80)
  • Regulated genes can be repressed and induced

The Jacob and Monad Operon Model
41
Regulation of bacterial genes
  • Response to nutrients in the environment
  • Example Inducible genes
  • Lactose operon

Figure 8.12a
42
Regulation of bacterial genes
  • Response to nutrients in the environment
  • Example Inducible genes
  • Lactose operon
  • Lac operon induction needs lactose AND
  • low glucose

43
Regulation of bacterial genes
  • Response to nutrients in the environment
  • Example Inducible genes
  • Lactose operon
  • Lac operon induction needs lactose AND
  • low glucose

Figure 8.13 - Overview
44
Regulation of bacterial genes
  • Response to nutrients in the environment
  • Example Repressible genes
  • Tryptophan operon

Figure 8.12b
45
Regulation of bacterial genes
  • 2. Response to accumulation of metabolic
    products
  • Example Feedback inhibition
  • Regulation of enzymes to synthesize
  • threonine

46
Regulation of bacterial genes
  • 3. Quorum sensing of the environment
  • QS ability of bacteria to communicate and
    coordinate
  • behavior (through gene expression) by release of
    signaling
  • molecules (phermones)

47
Regulation of bacterial genes
  • 3. Quorum sensing of the environment
  • Example Biofilm genes
  • Gram negative
  • homoserine lactones (HSL), a signaling
    molecule, results in loss of flagella,
    induction of virulence genes
  • Gram positives peptides

48
Regulation of bacterial genes
  • 3. Quorum sensing of the environment
  • Example Sporulation genes- low nutrients and QS
    induce
  • sporuation genes and shut down competence
    genes

Bacillus subtilis sporulation
49
Regulation of bacterial genes
  • 3. Quorum sensing of the environment
  • Example Virulence genes- P. aeruginosa uses QS
    to sense
  • high cell density and activate virulence genes
    to cause
  • disease

50
If you inoculate a flask of TSB with a single E.
coli, after 24 hours will all the cells be
identical? Why or why not?
51
Mutations base pair substitutions
  • 1. Missense and nonsense
  • mutations
  • Effect on protein
  • No change, altered function, loss of function
  • Examples
  • Sickle cell disease (mis)
  • Cystic fibrosis (non)
  • 2. Silent mutations
  • Effect on protein
  • No change in protein

52
Mutations frameshifts
  • 3. Frameshift mutations
  • Effect on protein
  • Many possible outcomes
  • Rarely no change
  • Examples (trinucleotide repeat diseases)
  • Fragile X Syndrome
  • Huntingtons Disease

53
Mutagens
54
Base analogs
Figure 8.18 - Overview
55
Mutations radiation
Figure 7.5
56
Mutations UV pyrimidine dimers
Figure 7.5
57
Mutations UV pyrimidine dimers
  • Solutions
  • Photoreactivation
  • - Photolyases

58
Mutations UV pyrimidine dimers
  • Solutions
  • Photoreactivation
  • - Photolyases remove
  • dimers
  • Nucleotide excision repair
  • - Can repair other damage

Figure 7.5
59
Mutations UV pyrimidine dimers
  • Solutions
  • Photoreactivation
  • - Photolyases remove
  • dimers
  • Nucleotide excision repair
  • - Can repair other damage
  • When all else fails
  • SOS! repair system
  • - Slows cell division,
  • but results in mutations

60
Mutations ionizing radiation
61
Independent Study
  • 1. Review material!
  • 2. Preview the mechanisms of genetic transfer in
    microbes
  • transformation, conjugation and transduction.
    (see Figures 8.24, 8.26,
  • and 8.27)
  • Print out APO-2 Have you thanked a microbe
    today?
  • Print out information for student presentations
  • - In Student Presentation folder
  • 1. List of individual assignments
  • 2. Presentation guidelines
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