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Microbiology > MICROBIAL GENETICS > Flashcards

MICROBIAL GENETICS Flashcards

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Biology 101 flashcards
1
Q

Transformation Griffith

A

First observed in 1928 by F. Griffith
Griffith was attempting to develop pneumococcal vaccine

Observation of transformation was incidental finding
Coined term “transforming factor”

Transformation was repeated in many experimental systems

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2
Q

Avery, Macleod and; McCarty

A

transforming factor as DNA

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3
Q

1980s technology-based transformation

A

becomes a workhorse of genetic engineering
Now one of the most commonly used methods for producing cloned DNA, and recombinant proteins
Summary diagram of principle follows

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4
Q

Transformation

A
  • Net result is acquisition of new phenotype resulting from introduction of new DNA sequences
  • Cell has undergone genetic recombination as a result of uptake of DNA from environmental solutions
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5
Q

Transformation Steps

A
  1. Recipient cell takes up Donor DNA
  2. Donor DNA aligns with complementary bases
  3. Recombination occurs between donor DNA and recipient DNA
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6
Q

Conjugation

A

-Bacterial sexual reproduction
-One cell acts as conjugation initiator
-DNA donor; F+ cell; male
-Carries F plasmid; produces F pili
-One cell acts as conjugation recipient
-DNA recipient; F- cell; female
-May be converted to F+ if conjugation is
completed
-Conjugation may cross species boundaries
-Hfr strains of bacteria have F plasmid genes integrated into main bacterial genetic element

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7
Q

Transduction

A

-DNA is transferred from a donor to a recipient cell via a bacteriophage mediator
-Bacteriophages are viruses that infect bacteria
-“normal” bacteriophage life cycle was elucidated
by Hershey & Chase (Waring blender experiment)
in 1952

    -Infection of host cells is initiated by “injection” of 
     viral DNA (only) into cell
              -Viral DNA then directs synthesis of new 
              viruses
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8
Q

Two major types Transduction

A

There are two major types of transduction

  • Generalized
    - associated with lytic cycle of bacteriophage
    - any gene of host cell can be transferred
    - bacterial dna combined w the phage goes into a new recipient cell

-Specialized
- associated with lysogenic cycle of bacteriophage
-only genes adjacent to prophage integration site
can be transferred

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9
Q

Transduction by a bacteriophage when bacteria dna is also is transferred

A
  1. A phage infects the donor bacterial cell
  2. Phage DNA and proteins are made, and the bacterial chromosome is broken into pieces
  3. Occasionally during phase assembly, pieces of bacterial DNA are packaged in a phage capsid. Then the donor cell lyses (opens up) and release phage particles containing bacterial DNA.
  4. A phage carrying bacterial DNA infects a new host cell, the recipient cell
  5. Recombination can occur, producing a recombinant cell with a genotype different from both the donor and recipients cell
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10
Q

Plasmids

A
  • Usually self replicating
  • Carry genes that confer unusual abilities
    - Toxin production
    - Resistance
    - Bacteriocins
  • Several types
    • Conjugative
    • Resistance
    • Dissimilative
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11
Q

Transposons

A
  • Discovered in the 1950s by Barbara McClintock
  • Small 0.7- 40 kb linear DNA segments
  • Contain transposase gene that allows them to move autonomously
  • Contain insertion sequences used when they move from one region of DNA to another
  • May also contain resistance, toxin or other genes that confer extraordinary abilities to the organisms that acquire them
  • Can easily cross species boundaries
  • Transposons can be incorporated into various types of plasmids
  • Confer extra mobility to the plasmids that carry them
  • May be very important evolutionary agents, as they exist in all cells, not just bacteria
  • Estimated frequency of transposition is comparable to spontaneous mutation rate in bacteria 10-5 to 10-7 per generation
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12
Q

Transposons

A
  • Can easily cross species boundaries
  • Transposons can be incorporated into various types of plasmids
  • Confer extra mobility to the plasmids that carry them
  • May be very important evolutionary agents, as they exist in all cells, not just bacteria
  • Estimated frequency of transposition is comparable to spontaneous mutation rate in bacteria 10-5 to 10-7 per generation
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13
Q

Transposons Steps

A
  • insertion sequence contain only a gene that codes for an enzyme transposase, which catalyzes the cutting and resealing of DNA that occurs in transposition and recognition sites
    • recognition sites are short inverted repeated sequences of DNA that the enzyme recognizes as recombination sites between the transposons and the chromosomes
  1. Transposase cuts DNA leaving sticky ends
  • complete transposons also carry other genes not connected with the transposition process.
    • bacterial transposons may contain genes for enterotoxin or for antibiotic resistance. Plasmids such as R factors are frequently made up of transposons
  1. Sticky ends of transposons and target DNA anneal
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14
Q

Regulation of Gene Expression (Eukaryote)

A

Eukaryotes

  • Goals
    - Embryonic development
    - Differentiation of tissues
    - Maturation and senescence
    - tumorigenesis
  • Multiple regulatory levels
    - Transcriptional
    - Post-transcriptional processing
    - Translational (free ribosomes vs. rough er)
    - Post-translational
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15
Q

Regulation of Gene Expression (Prokaryotes)

A 
Prokaryotes
-Goals
    -Conservation of biochemical and bioenergetic 
    resources
    -Cell division
    -Endospore formatio

-Because the key goal of regulation of gene expression
in prokaryotes is conservation of biochemical and
bioenergetic resources, regulation occurs at the
tightest level possible–transcriptional level

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16
Q

Two Classes of Prokaryotic Genes

A

Two Classes of Prokaryotic Genes

-Constitutively expressed–genes whose products are in constant use are not regulated; these genes are “always on” (transcribed & translated)
-Genes for major catabolic pathways such as
glycolysis
-Genes whose products regulate expression of other
genes

-Regulated genes–genes grouped into regulatory units within genome are found in OPERONS

17
Q

General Features of Operons

A

-Linear sequences of regulatory and structural genes

  • Regulatory sequences
    - Operator: binding site for operon regulatory protein
    - Promoter: RNA polymerase binding site

-Structural sequences: mRNA encoding region of operon includes a small number of genes whose products are functionally related

18
Q

Operon Regulation

A

-Genes for regulatory proteins are constitutively expressed
-Regulatory proteins bind to operator sequence
-Regulatory proteins are allosterically regulated by
either an inducer or a corepressor

-Two major classes of operons
-Inducible: normally “off,” use repressor protein &
inducer
-Repressible: normally “on,” use repressor protein &
corepressor

19
Q

Lactose Operon

A

-Mapped by Jacob & Monod in 1961
-Three structural genes encode enzymes used in the catabolism of lactose
--galactosidase: splits lactose to glucose &
galactose (lac Z)
-Permease: used in lactose transport (lac Y)
-Transacetylase: used in catabolism of other
disaccharides (lac A)

20
Q

Inducible Operon

A
  • Normally repressed; repressor protein is always available and bound to operator
  • Induction of operon occurs in response to high levels of lactose and low levels of glucose (glucose depletion leads to rise in cAMP levels); Lactose is converted to allolactose which then binds to repressor protein causing tertiary structure change that prevents repressor from binding to operator
  • RNA polymerase is now able to bind to promoter; transcription and translation of lac Z, Y, and A genes now freely occurs
21
Q

An Inducible Operon steps

A
  1. Structure of the operon
    • the operon consists of the promoter (p) and operator (o) sites and structural genes that code for the protein. The operon is regulated by the product of the regulatory gene
  2. Repressor active, operon off
    • The repressor protein binds with the operator, preventing transcription from the operon
  3. Repressor inactive, operon on
    • When the inducer allolactose binds to the repressor protein, the inactivated repressor can no longer block transcription. The structural genes are transcribes, ultimately resulting in the production of the enzymes needed for lactose catabolism
22
Q

Lac Operon (lactose present, glucose scarce)

A

if glucose is scarce, the high level of cAMP activates CAP, and the lac operon produces large amounts of mRNA for lactose digestion

23
Q

Lac Operon (lactose present, glucose present)

A

when glucose is present, cAMP is scarce, and CAP is unable to stimulate transcription

24
Q

Tryptophan (operon on)

A

an repressible operon.

  1. Structure of the operon.
    - the operon consists of the promoter (p) and operator (o) sites and structural genes that code for the protein. The operon is regulated by the product of the regulatory gene (I)
  2. Regressor inactive, operon
    - The repressor is inactive, and transcription and translation proceed, leading to the synthesis of tryptophan
  3. Repressor active, operon off.
    - When the corepressor tyrptohan binds to the repressor protein, the activated repressor bind with the operator, preventing transcription from the operon.
25
Q

Tryptophan (operon off)

A
  1. Structure of the operon.
    - The operon consists of the promoter (p) and operator (0) sites and structural genes that code for the protein. The operon is regulated by the product of the regulatory gene (i)
  2. Repressor inactive, operon on
    - The repressor is inactive, and transcription and translation proceed, leading to the synthesis of tryptophan
  3. Repressor active, operon off
    - when the corepressor typtophan binds to the repressor protein, the activated repressor binds with the operator, preventing transcription from the operon
26
Q

Transduction lytic cycle

A
  1. Phage attaches to host cell and injects DNA
  2. Phage DNA circularize and enters lytic cycle (Phage directs synthesis of viral components by the host cell)
    • mixes its dna into caspid
  3. New Phage DNA and proteins are synthesized and assembled into virions
  4. Cell lyses, releasing phage virions
27
Q

Transduction lysogenic cycle

A
  1. prophage exists in glactose-using host
  2. Phage genome separates carrying with it the adjacent gal gene from the host
  3. Phage matures and cell lyses releasing phage carrying gal gene
  4. Phage infects a cell that cannot utilize glactose (lacking gal gene)
  5. Along with the prophage, the bacterial gal gene becomes integrated into the new hosts DNA
  6. lysogenic cell can now metabolize glactose
28
Q

How does conjugation differ from transformation?

A
  • conjugation requires direct cell to cell contact
  • the conjugating cells must generally be of opposite mating type; donor cells must carry the plasmid, and recipient cells usually do not
29
Q

HFR cell conjugation

A
  • in some cells carrying F factors, the factor inegrates into the chromosome, converting the F+ cell to an Hfr (High frequency of recombination) cell.
  • When conjugation occurs between an Hfr cell and F- cell, the Hfr cell’s chromosome with its integrated F factor replicates, and a parental strand of the chromosome is transferred to the recipient cell.
30
Q

Conjugative Plasmid

A

-carries genes for sex pili and for the transfer of the plasmid to another cell

31
Q

Dissimilation plasmids

A
  • code for enzymes that trigger the catabolism of certain unusual sugars and hydrocarbons
  • helps them survive in diverse and challenging environments because of their ability to degrade and detoxify a variety of unusual compounds
32
Q

Resistance Plasmids

A

resistance to antibiotics, heavy metals, or cellular toxins

Microbiology (7 decks)

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