 View the video
“Microbial Evolution” and respond to student worksheet.
 Lecture: Species
Concept and Evolutionary
Domains. Respond to handout questions.

• http://www.youtube.com/watch?v=XawzIjX72U0
• http://www.youtube.com/watch?v=YPgxEl9jzRU&feature=relmfu
• http://www.youtube.com/watch?v=aF5sLLLalm8&feature=relmfu
• http://www.youtube.com/watch?v=vghlsa7oD_8&feature=relmfu
• 4 parts of video

• What is a species?
• A species is defined as a population that can naturally interbreed and produce fertile offspring, and that is reproductively isolated from other species.
• Right!
• Well, maybe not……..

• A bacterial species is a prokaryote whose 16S ribosomal RNA sequence differs by no more that 3%.
• http://www.microbeworld.org/caree rs/tools-of-the-trade/genetic-toolsand-techniques/16s-rrna
• 16S rRNA is part of the small subunit
(SSU) of the ribosome; used to classify prokaryotes.

• Prokaryotes do not fit the biological species concept because they
- Are haploid and reproduce asexually.
- Cannot produce “fertile offspring” like plants and animals can.
• In microbiology, evolutionary (molecular)chronometers measure evolutionary change.
- Chronometers are nucleotide or amino acid sequences that are homologous in different species.
- The greater the number of differences in the sequence the more distantly related the two species are.

• Molecular Chronometers
• Chronometer must be present in all groups being classified and it must be functionally homologous (not many sequence differences).
• The following genes and proteins are most frequently used to classify bacteria.
- ribosomal RNA
- ATPase proteins (synthesize ATP) 
- RecA (enzyme facilitates genetic recombination)
- Certain translation proteins.

• Ribosomal RNA is the most widely used chronometer for identifying bacterial species :
- It is relatively large.
- Universally distributed
- Has many nucleotide sequences that are conserved.

• Bacteria
• Prokaryotic organisms
• At least 40 phyla
• Have little in common phenotypically
• In terms of phylogenetics, most are genetically related.

• Archaea
• Prokaryotic organisms
• 4 phyla in this domain
• Contain extremophiles
- Hyperthermophiles: live in high temperatures.
- Methanogenic : produce methane.
- Extreme halophiles: live in high salt environments.

• Eukarya
• rRNA phylogeny based on 18S rRNA.
• Four kingdoms: Protista, Fungi,
Plant, and Animal.
• Range from single cell to complex multi-cell organisms.
• Rapid diversification of Eukarya was tied to changes in oxygen levels on earth.

• Questions for Speciation and Microbial Domains
• How is a bacterial species defined?
• What is an evolutionary (molecular) chronometer?
• What are 4 commonly used molecular chronometers?
• Why is 16S rRNA used to define bacterial species?
• What are the 3 domains in which microorganisms are found?
• Which domains contain prokaryotic organisms?
• Define hyperthermophiles, methangenic bacteria, and halophiles.
• What 4 kingdoms are contained in Eukarya

• Working microbiologists use phenotypic commonality in identifying bacteria. Most frequently these phenotypes are:
• Cell shape
• Cell wall structure
• Cell respiration
• Growth factors
• Colony morphology

• Cell Shape
• There are 4 bacterial shapes:
- Cocci (coccus s.) or round
- Bacilli (bacillus s.) or rod shaped
- Spirillum or cork screw shaped
- Filamentous or like jelly beans in straw

• Cocci
• Round shape
• Examples
-Staphlococcus 
(in clusters)
-Streptococcus
(in chains) 

• Bacilli
• Rod shaped
• Examples
- Bacillus anthracis
(agent in anthrax)

- Escherichia coli 
(used in biotechnology)

• Spirillum
• Cork screw shape
• Example
- Treponema pallidum
(agent of syphilis)

• Filamentous
• Jelly beans in straw
• Example
-Leptothris discophora (aquatic bacteria uses iron the way we use oxygen).

• Composition of Cell Walls
• Difference in cell wall structure becomes clear when a technique called the Gram stain is used.
• Bacteria on a glass slide are stained first with a purple dye; the slide is rinsed with ethanol, and then a red counter stain is applied.
• If bacteria remain purple = Gram positive.
• If bacteria turn red = Gram negative. http://www.youtube.com/watch?v=Qk2OjqatCqc&feature= related

• Gram positive cocci
• Gram negative rods

• Gram + organisms have a cell membrane and a cell wall made of peptidoglycan which gives the cell structure.

• Gram – organisms have a cell membrane and a cell wall with a thinner layer of peptidoglycan plus an outside layer of lipopolysaccharides. The lipopolysaccharides make Gram – organisms more virulent.

• Cell Respiration
• There are 3 types of cell respiration( synthesis of ATP):
- Aerobic: Use oxygen for cell respiration.
- Anaerobic: Cannot tolerate oxygen. Use fermentation
- Facultative anaerobes: Can use or not use oxygen depending on availability.

• Growth Factors
• Nutrient Source
- Heterotroph: Consume energy from outside source.
- Autotroph: Make and consume energy.
• Energy Source (Autotrophs)
- Chemoautotroph : Use chemicals as energy source.
- Phototrophs: Use light as energy source

• Colony morphology
• A single bacteria put onto a solid agar plate, if given sufficient nutrients, optimal temperature and pH, will multiply and form a colony.
• All members of the colony are genetically identical.
• Bacterial colonies of different species differ from one another.

• To identify a colony, the following basic elements are noted.
• Form- What is the basic shape of the colony?
• Elevation - What is the cross sectional shape of the colony?
• Margin - What is the magnified shape of the edge of the colony?
• Surface - How does the surface of the colony appear?
• Opacity – Is the colony translucent, transparent, iridescent?
• Chromogenesis – pigmentation.

• Form

• Elevation – Cross sectional shape

• Margin – Shape of colony edge

• Opacity – Clear, Opaque, Iridescent
Iridescent

• Chromogenesis – pigmentation

• Name and identify cell shapes
• Describe the similarity and difference between Gram positive and Gram negative cell walls.
• Explain the Gram stain procedure.
• Why is cell wall morphology important in clinical treatments?
• Define 3 types of cell respiration in bacteria.
• Describe 4 ways in which microbes produce and/or use energy?
• How are chemoautotrophs and photoautotrophs alike and different?
• What basic elements are noted in colony identification?
• Name and identify the colony forms.
• Name and identify the colony elevation.
• Name and identify the colony margin.
• Define chromogenesis.

 Webquest:
 Visit the 3 websites noted on your handout to learn about prokaryotic structures and function.
 Respond to all questions on the Webquest handout.
 Next read the Powerpoint slides on additional prokaryotic structures
 Respond to all questions on the handout.
 Class review and discussion

• http://www.cellsalive.com/cells/bactcell.htm#cytoplasm
• http://www.cellsalive.com/toc_micro.htm
• www.wisc-online.com/objects/ViewObject.aspx?ID=MBY901
 Complete your handout

• DNA in prokaryotes is found in the
- Nucleoid Region
- Plasmids
• A typical prokaryote has one chromosome containing most of the genes in the cell.
• A few species of Bacteria & Archaea contain two chromosomes.
• The DNA is a double stranded circular
molecule.
• Bacterial genomes contain from 500,000 base pairs to about 4 million base pairs, depending on the species..

 Plasmids
• Plasmids are DNA
• Exist and replicate separately from the chromosome.
• Most are circular, some are linear.
• Prokaryotes contain one or more plasmids.
• Range in size from 100 to 1,000 base pairs.
• Plasmid DNA can be exchanged among bacteria.
• Genes for antibiotic resistance are found on plasmids and one bacteria can transfer these genes to another bacteria.
• Bacterial plasmids play a role in recombinant DNA technology.

• The differences between a bacterial chromosome and a plasmid:
- Chromosomes carry many more genes than plasmids and the genes are essential to cellular function.
Essential genes are called
housekeeping genes.
- Plasmids carry far fewer genes and are expendable because the genes are not necessary for growth under all conditions.

• Restriction Endonucleases
• Are enzymes naturally found in bacteria.
• Have the ability to cut up the foreign viral
DNA if a virus is invading a bacterial cell.
The possibility of viral infection plummets.
• Can be thought of as a bacterial immune system because the role of restriction endonucleases is to protect the bacteria.
• Are of several types .
• Play a role in recombinant DNA technology

 Be able to define all prokaryotic structures and their functions; as listed in the Webquest.
 Be able to locate the structures on a diagram.
 Be able to answer the following questions from your Powerpoint slides
• Where is DNA found in bacteria?
• Describe the typical bacterial genome with respect to structure, location, and size.
• Describe a plasmid with respect to size, location, and function.
• Contrast bacterial chromosomal and plasmid DNA.
• How do restriction endonucleases work?

 Day 1: Hand washing and agar plate inoculation
 Day 2: Review plate streaking for colony isolation video, collect data, and streak plates for isolation.
 Day 3: Collect data, study colony morphology, and gram stain organisms.

Video Resources for Lab http://www.youtube.com/watch?v=PiWwnBbCrNs&feature=related
Pouring agar plates.
http://www.youtube.com/watch?v=zZ1NQau1wtw
Dilutions and spread plating http://www.youtube.com/watch?v=AaG3Pt3nwLQ&feature=relmfu
Streaking plates http://www.youtube.com/watch?v=tBmNitxvqyc
Aseptic transfer http://www.youtube.com/watch?v=SLkipIg4WRg
Making smears http://www.youtube.com/watch?v=-j97pZo5t4g&feature=related
Gram stains

 Lecture : E. coli and its use in biotechnology. Pathogenic
E.coli
 Reading: Pathogenic E. coli. Respond to handout questions.
 Case Study: Microbial Pie
 Activity: Track the Epidemic

• Escherichia coli
• Gram negative rod
• Facultative anaerobe
• Found in the intestines of warm blooded animals as normal flora.
• Benefits its host by producing vitamin K
• Reduces numbers of pathogenic bacteria in the intestine.

• E.coli is a hardy organism that is easy to culture and easy to manipulate in the lab.
• It is a model organism in biotechnology.
• Model organisms are extensively studied to understand biological phenomena and the information can be applied to other organisms.
• E.coli genome was one of the first to be sequenced in 1997

• Most economically robust area in biotechnology is production of human proteins.
• E.coli has played a major role in production of these proteins.
• Human genes for proteins can be cloned and inserted into plasmids in E.coli through recombinant DNA technology

• E.coli is then grown in large
bioreactors and it produces the protein of interest.
• Purification methods separate the target protein from the biological molecules in which it was produced.
• The proteins can then be used by humans.

The following proteins are manufactured via this technique:
 Insulin
For diabetes
 Human Growth Hormone
For growth hormone deficiency
 Factor VIII
For hemophilia
 Erythropoietin
For stimulation RBC growth

• Pathogenic vs. Non-pathogenic E.coli.
• Most E.coli strains live commensally in the intestines of warm blooded animals.
These strains are non-pathogenic .
• Non-pathogenic strains of E.coli strains are used in biotechnology research.
• Some E.coli strains are virulent and produce gastrointestinal disease. These strains are pathogenic .

• Causes of virulence
• Toxicity
- Ability to cause disease by a preformed toxin. Toxin inhibits host cell function and kills host cell.
• Invasiveness
- Ability of organism to grow in host cell tissue in such large numbers that pathogen inhibits host cell activity.

• E.coli virulence
• Due to an enterotoxin, a type of exotoxin.
• The enterotoxin is secreted by the bacteria and affects the cell membrane of intestinal cells.
• It makes the host cell membrane more permeable to chloride ions. As chloride enters the host cells, sodium and water leave the host cells.
• This causes diarrhea and abdominal pain.
• Virulent E.coli is acquired by eating contaminated food

• Where does nonpathogenic E.coli normally reside?
• What is a benefit of having E.coli as normal microbial flora?
• Explain the term “model organism”.
• Describe how E.coli is used to produce a protein such as insulin.
• Describe the 2 factors that can make E.coli a virulent organism.
• What are the 3 antigens used to type E.coli and where are these antigens located on the organism?
• What types of diseases can be caused by E.coli?
• What is the mode of transmission for gastrointestinal diseases?
• How do enterotoxins work?

• Lecture: Natural gene transfer and recombination.
• Whole class lecture: Gene transfer in prokaryotic organisms.
• Pantomime of gene transfer
• Case Study: Antibiotic resistance

• Bacteria pass on their DNA to the next generation asexually through binary fission.
• Many bacteria, however, have the capacity to physically exchange
DNA with other bacteria.
• There are 3 DNA exchange processes in
 Transformation
 Transduction
 Conjugation
• These 3 process are collectively referred to as lateral or horizontal gene transfer .

• Transformation
• Process by which free DNA is incorporated into a recipient cell and brings about genetic change.

• Do you remember the Griffith experiment?
http://science.jburroughs.org/mbahe/BioA/starranimations/c hapter8/videos_animations/griffith.html

• Transformation
• Bacterial cell is lysed. DNA pours out
• Bacterial chromosome breaks into 10 gene fragments genes.
• Other competent bacterial cells take up the DNA from the environment. Competency is genetically determined.
• DNA enters the cell & is escorted through the cytoplasm by competence specific proteins to prevent degradation.
• DNA is recombined (integrated) into bacterial chromosome.
• http://highered.mcgrawhill.com/sites/0072556781/student_view0/chapter13/animation_qui z_1.html

• Transformation in Biotechnology
• In biotechnology procedures, the term transformation has a slightly different meaning.
• E.coli are poorly transformed under natural conditions.
• If you treat the organism with calcium ions and chill it, it becomes easily transformed
(DNA from other source enters the cell).
• Transformation of this organism generally occurs in the plasmid.

• Transduction
• DNA is transferred from cell to cell by a virus.
• Virus can transfer host cell DNA along with its own genetic material.

• In transduction, any gene on a donor bacterial chromosome can be transferred to a recipient.
• A phage (virus for bacteria) enters the host cell & during a lytic infection enzymes responsible for packaging viral DNA sometimes package the host DNA accidentally.
• The resulting virus with a piece of the donor DNA is called a transducing particle.
• This transducing particle cannot go on to cause infection in a new cell. The DNA released is incorporated into a recipient bacterial cell chromosome.
• http://highered.mcgrawhill.com/sites/0072556781/student_view0/chapter13/animation_qui z_2.html

• Plasmids revisited
• Are genetic elements that replicate independently of the host chromosome.
• Are unessential, do not control vital cell functions.
• Are double stranded, mostly circular (some linear), structures with fewer genes than the bacterial chromosome.
• Of different types may be present in a cell and numbers of these types can vary.
• Called episomes can integrate into the bacterial chromosome.

• Types of plasmids
• F (fertility) plasmid - most studied, results in the expression of sex pili.
• R (resistance) plasmids - contain genes that can build a resistance against antibiotics
• Col plasmids - which contain genes that code for bacteriocins that can kill other bacteria.
• Degradative plasmids, which enable the digestion of unusual substances.
• Virulence plasmids- which turn the bacterium into a pathogen.

• Conjugation
• Process of genetic transfer that involves cell to cell contact.
• Conjugative plasmid uses this process to transfer a copy of itself to a new host.
• Process involves a donor cell and a recipient cell.

• Conjugation (using the F plasmid as an example)
• The F+ cell has the plasmid and is the donor.
• The F- cell is the recipient.
• F+ cell synthesizes a sex pillus.
• Sex pillus makes specific contact with the F- cell; pulling it toward the
F+ cell.
• The DNA (plasmid) is transferred from the F+ to the F- cell through the sex pillus.

• Depending on the species, the plasmid can be replicated first in the
F+ cell & then transmitted to F-. Other times, the 2 DNA strands are separated in F+ and one strand is transferred to F-. Both cells will then make a complimentary strand..
• The original F- cell turns into an F+ cell & can conjugate with other bacteria.
• Conjugative plasmids spread rapidly through populations much like infectious agents.
• If plasmids contain genes that offer a selective advantage (like an antibiotic resistance gene), this can ensure survival of that population.
• http://highered.mcgrawhill.com/sites/0072556781/student_view0/chapter13/animation_qui z_3.html

 First, work with a partner. Explain the processes of transformation, transduction, and conjugation. Next, have your partner explain the same to you.
 Next, create 6 groups. We will create pantomimes of horizontal gene transfer.
• 2 groups – Transformation
• 2 groups- Transduction
• 2 groups - Conjugation

• Be able to explain the process of transformation.
• Be able to explain the process of transduction.
• Be able to explain the process of conjugation.
( Use a good level of detail in your explanation and be sure to include appropriate vocabulary words)
• Describe plasmids.
• What are the different types of plasmids and their functions.

• Products of microbial biotechnology
• Read and familiarize yourself with the Powerpoint.
• Read each article on the website at the bottom of each slide and make a copy of it for your notes.
• Write a synopsis of each article

• Food Biotechnology - fermentation
• Scientists are currently working on ways to improve use of microorganisms in food production.
- Developing virus resistant organisms through recombinant DNA technology to prevent economic losses in the dairy industry.
- Developing bacteria to produce chemicals to kill contaminating organisms in food making processes.
- Produced a microbial enzyme used to make cheese. http://www.gmocompass.org/eng/grocery_shopping/processed_foods/29.dairy_prod
ucts_eggs_genetic_engineering.html

• Enzymes, Antibiotics, and Human Proteins.
• Recombinant DNA technology has enabled production of new enzymes, antibiotics, and human proteins from microbial fermentation.
• Prourokinase is an enzyme which helps heal wounds infected with
E.coli.
• New and novel antibiotics with two pathways for treatment are being developed.
• Tissue plasminogen activator, a protein which dissolves blood clots is being produced.
http://www.biotecharticles.com/Biotechnology-products-
Article/Biotechnology-in-the-Manufacturing-of-Detergents-159.html

• Fuels and Biopolymers
• Hydrogen power is a fuel of the future. Biotechnologists are looking at Clostridium species as generators of hydrogen.
• Plastics worldwide are polluters because they are not biodegradable.
Several organisms are being studied as producers of bioplastics.
These biodegradable plastics will have several applications in the industrial and medical fields. http://news.softpedia.com/news/Bacteria-Converts-Vegetables-to-
Bioplastic-167546.shtml

• Agriculture
• A Pseudomonas bacteria has been bioengineered with B.
thuringiensis toxin . The bacteria colonizes plants and acts as a biopesticide to kill insect larvae.
• Baculoviruses are used to contaminate plant material. Insects ingest the plant and develop a lethal viral infection Biotechnologists are working on ways to bioengineer the Baculovirus to enhance its ability as a biopesticide.
• http://archive.today/ANXQ

• Bioremediation
• Microorganisms with hydrocarbon oxidizing enzymes clean oil spills.
• Microorganisms are used in waste water treatment facilities to purify water.
• Bacteria are being studied which have the capacity to remove heavy metals such as arsenic, copper, tin, and mercury from the environment.
• http://freshscience.org.au/2003/aussie-arsenic-eating-bacteria-maysave-lives-and-clean-mines

• Discuss the relationship between renin and chymosin.
• What are the 4 types of enzyme based detergents? How do enzyme based detergents work vs. conventional detergents?
Why are enzyme based detergents preferable?
• Explain how bacteria convert fruits and vegetables into plastics?
• Describe how Baculoviruses act as an insecticide? What are the advantages and disadvantages of using this approach?
• Why does the bacterium NT-26 hold such promise in cleaning up mining areas?

• What are GMOs?
GMOs, or “genetically modified organisms,” are plants or animals that have been genetically engineered with DNA from bacteria, viruses or other plants and animals. These experimental combinations of genes from different species cannot occur in nature or in traditional crossbreeding.
• Visit the following websites for debate research
•
• http://classes.soe.ucsc.edu/cmpe080e/Spring05/projects/gmo/index.
htm http://www.csa.com/discoveryguides/gmfood/overview.php/editor.p
hp

• Refer to your handout for debate information and rubrics.
• You need to be able to:
- Provide 3 arguments in favor of GMOs.
- Provide 3 arguments against GMOs.
- Take a position for or against GMOs and support the argument.

• Reading: Fungal Biotechnology
• Lecture: Yeasts, Filamentous Fungi and Molds
• Video: The Biology of Fungi (16 min)

• Fungi – General Characteristics
• Fungi are composed of eukaryotic cells.
• Some are unicellular and some are multicellular.
• Habitats: Most are terrestrial and some are aquatic
• Energy : Fungi are heterotrophic decomposers. (A few are parasitic)
• Cell Walls: Resemble plants architecturally but are made of chitin not cellulose.
• Reproduction: Many reproduce asexually and sexually using spores.
• Recent molecular evidence suggests fungi are probably more closely related to animals than to plants or protists. http://www.fungionline.org.uk/

• Reading: Fungal Biotechnology
• Visit the following website: http://fungus.org.uk/nwfg/fungbiot.htm
Activity
• Provide 4 examples of why fungi are important in biotechnology and explain their use.

• There are 3 basic types of fungi
a. Unicellular fungi - Yeast
b. Filamentous fungi – Mold and fungi c. Macroscopic fungi –
Mushrooms
We will limit our discussion to the first two types

• Yeast
- 1,500 species of yeast
- not part of a single taxon.
• Cells
- typically spherical, oval, or cylindrical
- usually 3-4 microns in size
- most are unicellular
- some multicellular: a string of connected yeast cells connected by psuedohyphae.

• Yeast with pseudohyphae
• Pseudohyphae help yeast invade tissues.

• Energy
• Yeast flourish in environments where sugar is present.
• They are facultative aerobes; using aerobic cell respiration and fermentation.
• In a lab, yeast can be cultured with nutrient agar and grow colonies.

• Reproduction
• Yeasts generally reproduce
asexually by budding.
• http://www.youtube.com/wat ch?v=iOvrq6ssy2Y

• Reproduction
• Yeast can sexually reproduce by mating.
• Two different mating types fuse into a diploid cell.
• Diploid cell can bud to make additional diploid cells.
• Diploid cell undergoes meiosis and produces haploid cells called ascospores.
• Ascospores create new yeast cells.

• Yeast containing ascospores

• What are the general characteristics of fungi?
• Name the 3 general types of fungi and provide an example.
• Describe the following:
1. Structure of a yeast cell
2. Energy use in yeast
3. Asexual and sexual reproduction of yeast

• Filamentous Fungi
• Widespread in nature, usually seen on stale bread, cheese, or fruit.
• Called molds.

• Cell Structure
• A filament called a hypha (hyphae p,) grows from a single terminal cell.

• The hyphae grow together across a surface and form compact tufts called
mycelium.
• This compact mat represents many intertwined hyphae. 

• Cell Structure
• From the mycelium, hyphae grow upward.
• At the end of the vertical hyphae are
spores called conidia.

• Conidia are asexual spores and are often pigmented.
• http://bugs.bio.usyd.edu.au/learning/r esources/CAL/Microconcepts/Reprodu ction/fungiRepro.html

• Reproduction –
Asexual
• The function of the conidia is the dispersal of the fungus(via spores) to new habitats.
• When new conidia form they are white and eventually become pigmented.

• Reproduction - Sexual
• Fungi can reproduce sexually.
• An example is bread mold
Rhizopus.
• Hyphae called stolons of opposite mating types (+ & -) fuse to form a structure called gametangia
• Dipoid zygospore is formed.
• Zygospore produces sporandia which undergo meiosis and release haploid spores.

• Several yeasts, in particular Saccharomyces cerevisiae, have been widely used in biotechnology.
• S. cerevisiae is a simple eukaryotic cell, serving as a model organism for all eukaryotes.
• Fundamental cellular processes such as the cell cycle,, DNA replication, recombination, cell division, and metabolism have been studied.
• In 1996, S. cerevisiae was announced to be the first eukaryote to have its genome, consisting of 12 million base pairs, fully sequenced as part of the Genome project.
• S. cerevisiae as a model organism has improved our understanding of human disease genes.

• Describe the structure of a mold and the functions of each structure.
Include the terms fungal cell, hyphae, mycelium, and conidia in your description.
• Explain filamentous fungal asexual and sexual reproduction. Include the terms conidia, stolon, gametangia, zygospore, and sporandia in your description.
• What is the role of S. cerevisiae?

• http://www.youtube.com/watch?v=4NO299do_l4
• http://www.youtube.com/watch?v=Luxjo0AsbTY&feature=re lmfu

• Homework: Review and understand powerpoint and videos on virus structure, replication, and vectors.
• Class: Create 4 work groups and develop review questions on assigned slides
• Class: Present your slides and review questions to the class.
Write the review questions on the board; to be copied by students
• Work in groups of 4 to create a rap song involving virus content.
• Create and present a rap song about viruses.(See handout).

• Viruses as vectors
• Vector (in biotechnology) DNA that can be used to carry and replicate foreign DNA in biotechnology experiments.
• Viruses can serve as vectors. Genes of interest can be inserted into the viral genome and the genes of interest will replicate along with the virus.

• Viral vectors can be delivery system for gene therapy.
• http://www.edu365.cat/aulanet/comsoc/Lab_bio/simulacio ns/GeneTherapy/GeneTherapy.htm
• Viruses have potential as delivery systems in gene therapy because
• A. They naturally enter cells.
• B. They can integrate in the host cell genome.
• C. They are cell specific which would allow for targeted gene therapy .

• General Properties
• A minute particle containing nucleic acid, a protein coat, and sometimes other macromolecules.
• Can exist in extracellular or intracellular form.
• Extracellular –is metabolically inert.
• Intracellular – viral replication occurs.

• Genomes
• Viral genomes are very small (3 to 100 genes)and encode for those functions that they cannot adapt from their host.
• Viral genomes are categorized by the type of nucleic acid present.
a. Double stranded DNA b. Single stranded DNA c. Double stranded RNA d. Single stranded RNA e. Single stranded RNA that replicates with a DNA intermediate.
• Viral genomes can be linear or circular.

• Virus Structure
• Varies widely in size, shape, and chemical composition.
• Commonalities of structure
a. Nucleic acid (DNA or RNA)
b. Capsid- made of one to several proteins which surrounds nucleic acid.
c. Envelope –most animal viruses have an envelope. d. The envelope is composed of a phospholipid bilayer from the host and proteins which the virus makes.
e. Viruses without an envelope are called naked viruses.

• Enzymes
• Some viruses contain enzymes.
• Bacteriophages have lysozyme to make a small hole in bacterial cell wall.
• Retroviruses have reverse
transcriptase that transcribes DNA from their RNA.
• Viruses have enzymes because the cell would not be able to replicate the viruses with out them.

• Viral Replication
• Phases of replication process are a. Attachment b. Penetration c. Synthesis of nucleic acid &proteins d. Assembly e. Release

• Attachment
• Viruses - specific for the host cells they infect.
• Proteins on the outside of naked or enveloped virus interact with specific cell membrane receptors on host.
• If a cell membrane is altered, the virus cannot infect the cell; host resistance.
• However, viruses protein mutation enable viruses to interact with changed receptors.

• Penetration
• Three ways a virus can penetrate a cell membrane:
a. Membrane Fusion : Viral envelope and cell membrane fuse. Viral capsid alone brought into cytoplasm b. Entry Pore formation: Endocytosis of virus. Membrane removed in cytoplasm
c. Viral Penetration: The viral capsid is injected into the host cell's cytoplasm.

• Synthesis of nucleic acids and proteins - DNA viruses
• How viruses synthesize nucleic acids in the cell depends on the type of nucleic acid present in the virus.
• Double stranded DNA virus- Incorporates its
DNA into host genome & protein synthesis can begin.
• Single stranded DNA virus – Complimentary
DNA strand must be synthesized in the host because RNA polymerase requires double stranded DNA.

• Synthesis of nucleic acid and protein – RNA viruses
• RNA viruses need an RNA-dependent RNA-polymerase to replicate their
RNA.
• Cells do not have this enzyme.
• RNA viruses need to code for an RNA-dependent RNA polymerase.
• No viral proteins can be made until viral messenger RNA is available .
• The nature of the RNA in the virus affects its replication strategy.

• Synthesis of nucleic acids and proteins- RNA virus
• Single stranded RNA virus – There are 2 types of single stranded RNA viruses.
a. Plus-stranded RNA viruses -RNA is the same sense (direction) as mRNA and it functions as mRNA. This mRNA can be translated immediately upon infection of the host cell.
b. Negative-stranded RNA viruses - RNA is negative sense (complementary to mRNA) and must therefore be copied into the complementary plus-sense
mRNA before proteins can be made. The virus uses its own RNA polymerase to make the plus stranded m RNA.
• Double-stranded RNA virus - RNA is double stranded and can’t function as mRNA; Also need to package an RNA polymerase to make their mRNA after infection of the host cell.

• Synthesis of nucleic acids and proteins – retrovirus.
• The single strand of retrovirus RNA serves as a template to make a single strand of DNA with the virus’ enzyme reverse transcriptase.
• A complimentary DNA strand is made and the double stranded DNA is then a template for mRNA synthesis.
• http://highered.mcgrawhill.com/sites/0072495855/student_view0/chapter2
4/animation__hiv_replication.html

• Synthesis of nucleic acids and proteins
• Once mRNA is made proteins can by synthesized.
• Early proteins – are made first which are necessary for viral replication.
• Late proteins- are then synthesized such as the viral coat protein.

• Assembly
• Viruses self assemble in cells.
• Virus self-assembly within host cells has implications for the study of the origin of life, as it lends credence to the hypothesis that life could have started as self-assembling organic molecule

• All of the review questions you copied from the board.