Microbiology 3 Flashcards
Genetics
the science of heredity
Chromosomes
structures containing DNA that carry genes, microbes only have a single chromosome, we have a set of 2
Prokaryote chromosome
have a circular chromosome, genes are much more simple than eukaryotes
Eukaryote chromosome
have a linear chromosome (us), can preform gene splicing, genetics are very complex
Genes
the molecular unit of heredity
Alleles
different versions of genes, seen in eukaryotes only
Mutations
a source for different types of genes
DNA structure
double stranded helix, nucleic acid composed of nitrogenous bases
Nitrogenous bases
the base components of DNA and RNA, made of 5 carbon sugar and a phosphate group, they form the rungs of the structure
Nitrogenous base pairings DNA
C makes 3 hydrogen bonds with G, T makes 2 hydrogen bonds with A
Nitrogenous base pairings RNA
C makes 3 hydrogen bonds with G, U makes 2 hydrogen bonds with A
Genetic information transfer
DNA replication
Transcription
Translation
entire process takes place in the cytoplasm, all steps can occur at the same time, this can’t happen in eukaryotes
DNA replication
occurs before binary fission, must move from 5’ to 3’, is a semi-conservative process because each new DNA molecule contains one original strand and one new strand of DNA
DNA is anti-parallel
top strand is synthesized from 5’ to 3’, bottom strand synthesizes from 3’ to 5’ because it synthesizes in the opposite direction
Leading strand
DNA strand that continuously synthesizes
Lagging strand
DNA strand that synthesizes discontinuously
Origin
where DNA synthesis begins
Replication bubble
where the DNA strand opens up to be synthesized
Enzymes/molecules involved in DNA replication
DNA polymerase DNA ligase Helicase Single strand DNA binding proteins RNA primase Ribozyme
DNA polymerase
synthesizes DNA, can add nucleotides to the 3’ end only (OH), has a proof reading function to correct mutations
DNA ligase
covalently links the Okazaki fragments in lagging strand synthesis
Helicase
seperates the 2 strands of DNA and unwinds them
Single stranded DNA binding proteins
stabilize the strand of DNA, keeps the 2 strands separate by not allowing them to connect their hydrogen bonds
RNA primase
puts down RNA primer that is later removed and replaced with nucleotides, this allows us to have a 3’ hydroxyl for DNA polymerase
Ribozyme
RNA enzyme that removes introns and splices exons together, capable of acting as an enzyme
RNA synthesis
only one strand is copied
RNA polymerase
begins transcription when it binds to the DNA at the promoter site, synthesis continues until it reaches the terminator site on the DNA
Promoter sequence
indicate the start of a gene
RNA types (3)
rRNA
mRNA
tRNA
rRNA
forms integral part of ribosomes
Ribosomes
a minute particle consisting of RNA and associated proteins, cellular machinery for protein synthesis, bind mRNA and tRNA to build polypeptides and proteins, found in large numbers in cell ctoplasm
mRNA
carries coded information that must betranslated, ultimately results in a protein
tRNA
structural RNA, involved in protein synthesis
Important tRNA sites
amino acid binding site, anticodon
mRNA codons
there are 64 codons and only 20 amino acids, the code will be redundant
Genetic code
is redundant, universal or nearly universal, 64 codons, 61 are sense codons, 3 are non-sense codons
Sense codons
code for an amino acid
Non-sense codons
aka stop codons, you hit one about 5% of the time
AUG
is the start codon
Regulation of metabolism
80% of bacteria are not regulated
Constitutive
bacteria that are not regulated and are being produces all the time
Feedback inhibition
enzymatic, end product is threonine which goes back to enzyme 1 and shuts down the pathway through non-competitive inhibition
Genetic regulation of metabolism
uses operons, I gene is upstream from the operon and is always on
Mutation types
point mutation
frame shift
Point mutations
silent
missense
nonsense
Silent mutations
base substitution, has no effect on the organism
Missense mutations
coding for the wrong amino acid
Nonsense mutations
base substitution mutation, codes for a stop codon then the sequence is not completely done
Causes of mutations
spontaneous
induced
chemical mutations
radiation
Spontaneous mutations
arise during replication
Induced mutations
chemical mutagens
ex: acridine: frame shift, wedges into double helix causing a frame shift
Chemical mutations
Base analog
5-bromouracid is inserted into DNA instead of thymine, base pairs with Guanine
Radiation
causes adjacent pyrimidines to bond, transcription of mRNA stops at the gap
DNA repair
Light repair
Dark repair
uses DNA polymerase, ligase, endonucleases, and exonuclease
Light repair
light activates photolyases that break dimers
Dark repair
can occur with or without light, uses nucleotide excision repair
Ways to acquire mutation
Induced
spontaneous
Induced mutations
exposure to an antibiotic induced a change in an organism, mutations occur only in the presence of antibiotics
Spontaneous mutations
allows the organism to grow in an antibiotic, this selects for the resistant mutant, there will be large fluctuations in the number of resistant organisms per culture, a mutation can occur early or late in the incubation period
Fluctuation test
used to determine whether mutations were spontaneous or induced
Replica plating method
used to study mutations, sterile velveteen pad is imprinted on master plate, in the same orientation, the pad is used to inoculate an agar plate with the antibiotic
Conclusor
used to study mutations, bacteria on the antibiotic plate had resistance without exposure, this demonstrates the spontaneous nature of mutations
Ames test
used to screen chemicals for their mutagenic properties, uses histidine autotrophes of salmonella, upon exposure to mutadine, they have the ability to revert back to histidine synthesizing capability
Carcinogens
tend to be mutagens
Auxotroph
nutritionally deficient mutant
Resistance plasmids
aka R plasmids, resistance is not induced by antibiotics, resistant strains are selected for by antibiotic use
Genetic engineering
the direct manipulation of genes for practical purposes
Genetic engineering techniques
protoplast fusion, recombinant DNA cloning
Protoplast fusion
protoplasts of 2 strains can be mixed to allow for genetic recombination of desired characteristics
EX: slow growing, good producer of substance fuse when there is polyethylene glycol with a fast growing poor producer to get a fast growing good producer
Protoplast
organism with its cell wall enzymatically removed
Recombinant DNA
DNA from 2 different sources covalently linked to create a single DNA
If a plasmid is cut with the same restriction enzyme, the 2 DNAs will have compatible “sticky ends”, can covalently link the 2 DNA with DNA ligase, this makes recombinant DNA
Gene cloning
the production of multiple copies of a gene carrying pieces of DNA, recombinant plasmid is used to transform bacteria, recombinant bacteria are selected for using media with an antibiotic, clonal population of cells create multiple copies of the gene
Viruses
obligate intracellular parasites, can only replicate inside a host cell
Virus nucleoproteins
nucleic acid covered by a protein coat, viral genome may be either DNA or RNA
Viral components
nucleic acid core
capsid
envelope
Viral nucleocapsid
naked (no envelope)
Viral nucleic acid core
genome may be DNA or RNA, ss- or ds-, linear, circular or segmented
Viral capsid
protein coat that surrounds the genome, composed of capsomeres
Viral envelope
bilayer membrane spikes, glycoproteins attach to host receptor
Viral size
must be viewed with an EM, ribosome = 25-30nm
Viral shapes
Heilcal
Polyhedral
Complex capsid
Helical shape
capsid forms helix around genome
Polyhedral shape
capsid is many sided, most common
Icosahedron
20 triangular faces
Complex capsid shape
combination of helical and icosahedral shapes
Viral host range
the spectrum of hosts that a virus can infect
West Nile virus
a good example of a broad host range
Viral specificity
Virus is selective in the organisms it infects, the type of cells and disease it produces
What is used to classify viruses
based on type and structure of nucleic acid genome
DNA or RNA genome
Double or single stranded
linear, circular or segmented
Virus families
-viridae
are distinguished on the basis of nucleic acid type, capsid shape, presence of envelope and size
RNA virus chromosomal arrangements
is a single strand, viruses do not have both + and –
+ sense RNA viruses
during infection, RNA acts like mRNA and is translated
- sense RNA viruses
RNA acts as a template for the production of + sense RNA, must carry RNA polymerase with the virion
Rhabdoviridae
sense RNA virus, enveloped, helical, 70-180nm in size, virion contains an RNA dependent RNA polymerase
Rhabdoviridae Ex
Rabies virus
Double stranded RNA viruses
one family with the virion, has segmented dsRNA
RNA virus families
picornaviridae
retroviridae
rhabdoviridae
Picornaviridae
+ sense RNA virus, naked, polyhedral shape, 18-30nm in size, translated to produce an RNA , dependent RNA polymerase
Picornaviridae Ex
polio virus
Retroviridae
+ sense virus, retro transcription, enveloped, spherical, nm in size, virion contains 2 copies of the genome and the enzyme reverse transcriptase (makes DNA from RNA template)
Retroviridae Ex
provirus: before transcription, new DNA is incorporated into the host genome
DNA virus families
grouped on basis of DNA structure, only one family has ssDNA
Herpesviridae
linear dsDNA virus, enveloped, polyhedral, 120-200nm in size, viral dsDNA can exist as a provirus, causes latent infections
Latent infections
virus remains in the host for a long time, can still replicate
Viral, Bacteriophage and anvimal DNA virus replication steps
Adsorption Penetration Synthesis Maturation Release
Viral Adsorption
the attachment of viruses to host cells
Viral Penetration
entry into host cells
Viral synthesis
creation of new nucleic acid molecules, capsid proteins using the host’s metabolic matching
Viral maturation
assembly of these components into infectious virions
Viral release
departure of new virions, generally killing the cell
T-even bacteriophage
dsDNA, complex, naked, capsid head collar
T-even bacteriophage host Ex
E. Coli
Bacteriophage adsorption
specific proteins on the tail fibers bind to specific receptors on host cells
Bacteriophage penetration
Lysozyme weakens the cell wall, the tail sheath contracts, viral genome is “injected” from head into bacterial cell
Bacteriophage synthesis
phage directs host cell to make phage products, bacterial DNA is disrupted
Bacteriophage maturation
viral components are assembled into infectious virions
Bacteriophage release
lytic phage lyse the host cell and inject neighboring cells
Bacteriophage growth curve stages
Eclipse period
Latent period
Viral yield
Bacteriophage growth curve eclipse period
spans from penetration through synthesis
Bacteriophage growth curve latent period
spans from penetration to phage release
Bacteriophage growth curve viral yield
number of viruses per injected cell
Plaque assay
used to determine phage number
reported in pfu (plaque forming units)
Plaques
clear areas where phage has infected host and surrounding cells
Temperate bacteriophage
Lysogeny
Bacteriophage doesn’t kill host
Lysogenic conversion
prevents adsorption of similar phage and biosynthesis of prophage
Lysogenic prophage
produces proteins that repress viral replication
Lysogenic induction
spontaneous or induced excision of prophage resulting in lytic cycle
Animal DNA virus adsorption
enveloped viruses have spikes that bind receptors
Animal DNA virus penetration
Nucleic acid and capsid enter cell
uses uncoating
Animal DNA virus synthesis
Viral DNA genome is replicated and viral proteins are synthesized, viral proteins move to the nucleus where they combine with new viral DNA
Animal DNA virus maturation
the complete virion is assembled; enveloped viruses bud through a host membrane where viral lipids and glycoproteins are present
Animal DNA virus release
Budding of new virions does not necessarily kill the host cell
Transcription
occurs before we express a protein, is the synthesis of a complimentary strand of RNA from a DNA template
Translation
decodes the “language of nuclaic acids to proteins, occurs on the ribosome
Translation steps
Initiation
Elongation
Termination
Translation elongation steps
codon recognition
peptide bond formation
translation
Polyribosome
transcription and translation occurring at the same time
Genetic regulation of metabolism steps
enzyme induction
enzyme repression
Enzyme reperssion
uses lac operons and trpoperons
Operon consists of
Promoter sequence
operator
structural genes
Enzyme repression: Operator
binds to repressor
Enzyme repression: Structural genes
make a protein code for enzymes
Lac operon without lactose
repressor bound to operator
Lac operon with lactose
lactose is converted to allolactose
allolactose
the inducer, binds to repressor and inactivates it so it no longer binds to the operator
Trp operon
I gene makes an inactive repressor, default position is on
Trp operon Tryptophan
acts as a co-repressor, tyrp binds to the repressor and activates it
Trp + repressor
would bind to the operator and shut down transcription
Bacteriophage
a virus that infects bacteria, nucleic acid core is covered by a protein coat
Bacteriophage life cycle
2 possible outcomes:
lytic cycle
lysogenic cycle
Bacteriophage lytic cycle
characteristics of virulent phages, the cell is lysed releasing hundreds of bacteriophages
Bacteriophage lysogenic cycle
initiated by a temperate phage, phage is incorporated into bacterial chromosome and replicated with it
Emerging viruses
increase in viral disease
Emerging virus causes
previously endemic viruses can spread due to global warming
ex: delongue fever
tropical islands are farmed and contact viral vectors
ex: yellow fever
viral host range spread to other species, becomes a mutant virus not recognized by the immune system
ex: swine flu pandemic of 1918
Bacteriophage structural components
Genome
tail sheath
plate and tail fibers
Bacteriophage genome function
carries the genetic information necessary for replication of new phage particles
Bacteriophage tail sheath function
retracts so that the genome can move from the head into the host cell’s cytoplasm
Bacteriophage plate and tail fibers function
attach phage to specific receptor sites on the cell wall of a susceptible host bacterium
Prophage
Phage genome is incorporated into host nucleic acid
Lysogen
Bacterium and temperate phage
Uncoating
enzymes digest the protein coat releasing the DNA
RNA viruses examples
polio
HIV
both are + sense ssRNA
Polio adsorption
naked viruses have proteins that bind to complementary proteins on the host
Polio penetration/uncoating
most naked viruses enter the cell by endocytosis
Polio synthesis
mRNA and viral proteins are produced, + sense RNA acts as template to make - sense RNA, -sense RNA is the template for making many copies of the + sense RNA viral genome
Polio maturation
new virions are assembled in the cytoplasm
Polio release
kills the host cell
HIV adsorption
glycoprotein spikes in the envelope recognize protein receptors on the host cell surface
HIV penetration/uncoating
fusion with the host
HIV synthesis
2 copies of + sense RNA copied into ssDNA by reverse transcriptase, second strand of DNA is synthesized, dsDNA inserts into host cell genome as a provirus, provirus genes are transcribed and translated
HIV maturation
2 copies of + sense RNA are packed into each capsid
HIV release
mature HIV nucleocapsids bud from the plasma membrane
Culturing animal viruses
eggs cell culture primary cell culture diploid fibroblast strains continuous cell line cytopathic effects
Culturing-eggs
fertilized, intact eggs used to grow animal viruses, difficult to study cellular effects caused by viruses
Cell culture
animal cells are grown in monolayers, animal cells are treated with proteolytic enzymes, cells can be subcultured after growth
Primary cell culture
comes directly from the animal, cells usually divide a few times
Diploid fibroblast strains
immature cells that produce collagen, derived from fetal tissue, retain capacity for repeated cell division
Continuous cell line
will reproduce for extended number of cell divisions
ex: HeLa line = cervical cancer
Cytopathic effects
CPE
visable effect viral infection has on cells
ex: cells change shape, detatch from culture container
Virus effects
transformation
teratogenesis
Viral transformation
another CPE caused by viruses, the conversion of normal cells to malignant ones
Viral teratogenesis
induction of defects during embryonic development
Teratogen
drug or agent that causes defects, viruses can act as teratogens
ex: CMV, HSV and rubella
Virus like agents
viroids
prions
Viroids
infectious RNA particles smaller than a virus, no protein products are produced, disrupt host cell metabolism, cause lethal plant diseases
Prions
proteinaceous particles, proteins exist in normal form and prions proteins stick together and eventually kill cells
Tumor or neoplasm
localized accumulation of cells
HPV
human papillomavirus, causes cancer, dsDNA, exists as a provirus, production of excess viral replication protein causes uncontrolled cell growth
Cancer from a virus
15% of cancer comes from a virus, discovered in 1911
Oncogenes
a gene that causes uncontrolled cell growth
Proto-oncogenes
a normal gene that when under the control of a virus can act as an oncogene
