MCB Exam 2

What is meant by recombinant DNA technology? Describe the use of it in producing Insulin. What is the purpose of cloning?

rDNA technology: the insertion or modification of genes to produce desired proteins. Purpose of cloning: To have identical cells arising from one cell. Main goals 1) Eliminate undesirable phenotypes 2) Combine beneficial traits 3) Create organisms that can produce human product (Insulin)

Describe gene gun, microinjection.

Microinjection: Is a technique for introducing a solution of DNA into a cell using a fine micro-capillary pipette. Gene Gun: A way of introducing foreign DNA into plants cells..

Compare the general features of Bacteria, Archaea, and Eukarya.

Bacteria: Prokaryotic, contains peptidoglycan, has rRNA loop Archaea: Prokaryotic, cell wall varies in composition; contains no peptidoglycan, lacks rRNA Eukarya: Eukaryotic, cell wall varies in composition, but contains carbohydrates, lacks rRNA.

What is the order of virus life cycle? What occurs during adsorption, un-coating and assembly? Describe one step growth cycle. How one grows viruses in lab?

1. Attachment: virus attach to host cell by capsid2. Penetration: Inject virus genetic material like DNA into host cell3. Un-coating: Releasing capsid inside host cell to outside (not always done since capsid usually stays outside of cell)4. Synthesis/Replication: Virus makes viral proteins and nucleic acids (virus takes over host common center for protein and DNA synthesis and now makes copies of its own genes), host cell can't divide5. Assembly: Maturation and Packaging, self-assembly of virions6. Release: release of viruses out of the host cell-One-step growth cycle for viruses: Attach to the host and then penetrate and go inside the host, then the virus becomes invisible while biosynthesis and maturation is going on and can not be seen called the eclipse period, but then the virus releases all the replicated viruses at the same time and kills the host by lysis and now you can see all the new viruses (latent period is when penetrated till virus release)-Measure viral growth in animal cells: by using the plaque assay (dilutions of the virus are used to infect a cultured cell monolayer from an organism which has agar in the media to make it solid), if virus attacks the monolayer host then it will be concentrated in certain areas of cells and you can see black dots that are the plaques, The number of plaque (black dots) on the medium EQUALs the number of infectious virus particles present on plate and then multiple by dilution factor to get total.

List some chemical agents that can cause cancer (mutagens) and describe their mechanism of action.

Nitrous acid: causes adenine to bind with cytosine instead of thymine Nucleoside analog: incorporates into DNA in place of a normal base; causes mistakes in base pairing.

Define base analogs and explain how they may induce mutation.

Base analogues are molecules that can substitute for normal bases in nucleic acids. Usually, substitution of a base analogue will result in altered base pairings and structural changes that affect DNA replication and transcription of genes

List the differences between bacteria and viruses. Where in nature can viruses be found.

Bacteria have own reproductive machinery, they must have DNA, have cell wall Viruses must use host to reproduce, can have DNA or RNA, have a capsid that wraps itself around the nucleic acid

List scientific and agricultural applications of recombinant DNA.

Agricultural applications: Bt toxin > plants have toxin-producing gene, toxins kills insects that eat plants (Bt cotton, Bt corn) Herbicide resistance Suppression of genes-antisense DNA (improve shelf life) Nutrition-human proteins RoundUp (glyphosate) >plants have bacterial gene; allows use of herbicides on weeds without damaging crops. Scientific applications: genetic screening (southern blots used to screen parents and fetuses for inherited diseases caused by mutations); DNA fingerprinting; Gene therapy: replace defective and missing genes with normal ones

Define operons and explain their role in gene expression in bacteria. Give examples.

An operon is a group of genes that are transcribed together and controlled by one promoter. It is for regulation gene expression. Inducible operons: (Catabolic reactions, sugar metabolism controlled by inducible operons) they are usually off unless there is a substrate for them to metabolize. "Off" Repressible operons: control genes involved in synthesis (anabolic reactions) Tryptophan operon is an example "On" because they are synthesis processes, and they are off when amino acid comes from environment and the need to make them is no longer present.

Which of the three domains of microbes are prokaryotes and which are eukaryotes?

Bacteria and Archaea> Prokaryotes Eukarya> Eukaryotes

What is plaque and what is it used for? What is CPE?

CPE> Cytopathic effects: ability to center the cell. Ability to multiply within the cell and to evade host immune system. Ability to damage hose (visual damage either macroscopic or microscopic). Examples: cell lysis or plaque formation altering nuclear or cytoplasmic structures. Changes in cell membrane elasticity. Take control of host gene expression and metabolism. Inhibits host protein synthesis. Encourage host cell apoptosis, cell fusion.

List features of Chlamydia, spirochetes and mycoplasma.

Chlamydiae: obligate intracellular bacteria, causing eye infection, chlamydia. They have elementary body a small inactive use for transmission of disease. Spirochetes: causes syphilis and Lyme disease they move via axial filaments. Mycoplasma: bacteria with no cell wall> causes mild pneumonia.

Define chromosome, gene, genome.

Chromosome- structures containing DNA that physically carry hereditary information; the chromosomes contain genes. Gene- segments of DNA that encode functional products, usually protein Genome- all the genetic information in a cell

Define and explain: Complementary strands, anti-parallel, semiconservative replication, double helix.

Complementary strands: strands of DNA that are paired up AT & CG Anti-parallel: parallel but moving or oriented in opposite directions(5' to 3' and 3' to 5') Semiconservative replication: describes the mechanism by which DNA is replicated in all known cells. one strand is original from parent, one strand is new (1/2 new, 1/2 from parent) double helix: a pair of parallel helices intertwined about a common axis, especially that in the structure of the DNA molecule.

Define frame shift mutation, nonsense mutation.

Missense mutation: base substitution results in change in an amino acid Nonsense mutation: base substitution results in a nonsense (stop) codon Frameshift mutation: Insertion or deletion of one or more nucleotide pairs. Shifts the translational "reading frame"

Describe the use of human microbiome in crime scene investigation.

DNA fingerprinting (RFLP) is used to identify pathogens. This can be used to track infectious diseases. Take DNA cut it into pieces>gel electrophoresis> then take sample from father and child and compare. Can be used in forensic science. You should notice the same pattern in the gel.

Define function: DNA gyrase, DNA ligase, DNA polymerase, RNA polymerase, helicase.

DNA gyrase: Relaxes supercoiling ahead of the replication fork DNA Ligase: Makes covalent bonds to join DNA strands; Okazaki fragments, and new segments in excision repair. DNA polymerase: Synthesize DNA; proofread and facilitate repair of DNA. RNA polymerase: Synthesizes RNA from DNA. Helicase: Unwinds double-stranded DNA.

What is the genetic material for viruses?

DNA or RNA, either single or double stranded.

Define Central Dogma of Molecular Biology.

DNA>mRNA>Protein>Function. This describes how, typically, DNA is transcribed to messenger RNA, which, in turn, is translated into proteins that carry out vital cellular functions. "Flow of genetic information"

What are important points discussed in "Big Picture-Genetics"

Genetics is the study of genes: how they are replicated, expressed, and passed on from one generation to another. Mutations introduce change into this process, which can be frameshift or base substitution. Genes in operons can be inducible "off" with repressor bound to DNA or repressible "on" with repressor not bound to DNA. Alteration of bacterial genes and/or gene expression may cause disease, prevent disease treatment, or be manipulated for human benefit.

What are the products of replication, transcription, translation and reverse transcription? What are the differences between DNA polymerase and RNA polymerase? What is the significance of reverse transcriptase? What is the source of reverse transcriptase. What does it do?

In replication, one strand of DNA serves as template for another, thus the products are 2 identical copies of DNA. Transcription products: RNA Translation products: proteins DNA Polymerase synthesizes a DNA strand and used in DNA replication while RNA Polymerase is used during transcription to synthesize the MRNA strand. In contrast with the DNA polymerase, RNA polymerases do not necessarily require the so called primer to start the process and they actually have no proofreading systems. makes DNA from RNA-unique to some viruses (HIV)-makes DNA from RNA

How influenza virus may cross the species barrier and move from animals to humans

Influenza A viruses are found in many different animals, including birds, pigs, whales, horses, and seals. Sometimes influenza A viruses seen in one species can cross over and cause illness in another species

How different types of radiation may cause mutation.

Ionizing radiation (X rays and gamma rays) causes the formation of ions that can oxidize nucleotides and break the deoxyribose-phosphate backbone UV radiation causes thymine dimers Repair (UV only)> photolyases (separate thymine dimers) and nucleotide excision repair (enzymes cut out incorrect bases and fill in correct bases)

What is the difference between the two forms of radiation?

Ionizing radiation> more serious mutation. DNA is actually broken here. You cannot repair broken DNA UV radiation> fuses two T together (thymine dimers). You can fix the thymine dimer.

List forms of fungal diseases.

Mycosis: fungal infection Systemic mycoses: deep within the body Subcutaneous mycoses: beneath the skin Cutaneous mycoses: affect hair, skin and nails Superficial mycoses: localized Opportunistic mycoses: fungi harmless in normal habitat but pathogenic in a compromised host

Define and compare latent viral infections and persistent viral infections.

Latent viral infections: No virus is produced, no host dies, virus becomes part of the host chromosome. The virus gets in an attacks the host but does not kill the host, rather it remains with the host. Persistent: Sometimes called chronic infection> some virus produced some host die but slow and over a long period of time

Define lysogeny and describe its role in passing disease causing genes from one bacterium to another. What other genes may be exchanged using lysogenic viruses.

Lysogeny: phage remains latent, not active, not multiplying and does not kill the host. They insert DNA into host cells DNA at which point becomes known as prophage. When the host cell replicates its chromosome, it also replicates prophage DNA> phage conversion (host exhibits new properties)

Important points discussed in "Clinical Focus-Norovirus Outbreak"

Noroviruses are the most common cause of outbreaks of acute gastroenteritis worldwide. Annually, norovirus causes 20 million cases of gastroenteritis. During 2015, 316 norovirus outbreaks in the United States were reported

What is meant by obligate intracellular pathogens? Which group of infectious agents are included?

Obligatory intracellular pathogens: require living host cells to multiply. They contain DNA or RNA, a protein coat, no ribosomes, no ATP generating mechanism. Every virus must have a protein capsid and DNA/RNA.

How viruses may transform a normal cell into a tumor cell?

Oncogenes transform normal cells into cancerous cells. The oncogenic viruses become integrated into the host cell's DNA and induce tumors.

What is the role of plasmids in horizontal gene transfer?

Plasmids are self-replicating circular pieces of DNA carry genes that can cause diseases and antibiotic resistant genes. They may code for proteins that enhance the pathogenicity of bacterium

Name different shapes of viruses? Which is unique to viruses?

Polyhedral virus: multiple faces Helical Virus: Ebola virus is an example. Complex: bacteriophage Some have envelope (Flu virus)

Importance of cyanobacteria/ algae

Primary producers that fix CO2 through photosynthesis to produce oxygen.

Where in eukaryotes and prokaryotes can one find DNA?

Prokaryotes DNA is found in: circular or linear chromosomes in supercoil in nucleoid, plasmid Eukaryotes DNA is found in: linear chromosomes in nucleus, mitochondria and chloroplast in plants

Compare general features of prokaryotic and eukaryotic cells.

Prokaryotic structures lack membrane enclosed organelles and a nucleus, but usually contain a cell wall. Eukaryotic structures contain membrane enclosed organelles and a nucleus, but lack a cell wall. Prokaryotic cell: DNA circular, ribosomes, binary fission Eukaryotic cell: linear DNA, ribosome, mitosis

Describe general features of Protozoa, Fungi, Viruses.

Protozoa: a catchall kingdom for a variety of organisms; autotrophic and heterotrophic (clades based on rRNA) Fungi: chemoheterotrophic; unicellular or multicellular; cell walls of chitin; develop from spores (decomposers) Plantae: multicellular; cellulose well walls Archaea: grow in extreme conditions and are very beneficial to the planet. Viruses: submicroscopic, obligate intracellular parasites with host specificity. They can infect plants, fungi, bacteria, Archae and are acellular.

What are the general features of Protozoa? Compare cyst and trophozoite.

Protozoa: unicellular eukaryotes, inhabit water and soil, complex life cycles. Cysts: for transmission and inactive, resistant to elements Trophozoite: growing, multiplying, inside and grows

Name different RNA molecules. What is the role of each molecule? Which one contains Codon, anticodon.

RNA (ribonucleic acid): single stranded nucleotide, 5 Carbon ribose sugar, contains Uracil (U) instead of Thymine (T). rRNA: integral part of ribosomes tRNA: transports amino acids during protein synthesis. mRNA: carries coded information from DNA to ribosome. mRNA has codons > ribosome reads the codon> tRNA carries anti-codon to ribosome >ribosome makes a protein.

What is meant by pre-transcriptional control?

Repression and induction regulate the transcription of mRNA. These mechanisms control the formation and amounts of enzymes in the cell.

Name the tools used to make a recombinant DNA. Define gene library and protoplast fussion.

Restriction enzymes: cut specific sequences of DNA. Vectors: carry new DNA to desired cells (usually plasmids or viruses DNA is cut and can be inserted into a cell via Transformation> cells take up DNA from the surrounding environment Electroporation> electrical current forms pores in cell membranes Protoplast fusion> removing cell walls from two bacteria allows them to fuse. Gene library: Collections of clones containing different DNA fragments.

Explain how a technique called RFLP works in analyzing DNA.

Restriction fragment length polymorphisms are different lengths of DNA restriction fragments from different individuals of same species because of the deletion or addition of DNA between sites where restriction enzyme cuts.-Used to determine ancestry of individual and identify DNA from specific individual, determine the location of genes causing genetic diseases and identifies new inserted genes or DNA sequences. Take DNA cut it into pieces>gel electrophoresis> then take sample from father and child and compare. Can be used in forensic science.

What are ribosomes made of? Prokaryotes and Eukaryotes have different ribosomes? What is the significance of the difference?

Ribosomes are made of rRNA and proteins.- Difference between prokaryotes and eukaryotes ribosomes: size and composition, prokaryote ribosomes are a target for antibiotics. eukaryotes have bigger and denser ribosomes- Significane of this difference: antibiotics (Tetracycline and Erythromycine) won't kill eukaryote ribosomes just prokaryote ribosomes so kill bacteria inside us

What is the goal/purpose of Genome Project?

The human genome project sequenced the entire human genome.

Economic impact of fungi

Saccharomyces cerevisiae: bread, wine, hepatitis B vaccine Trichoderma: cellulase Taxomyces: taxol Entomophaga: biocontrol Coniothyrium minivans: kills fungi on crops Corn>alcohol>fuel.

What is the function of microRNAs (miRNAs)?

Small, single stranded RNA that prevent translation of a complementary mRNA. They inhibit protein production in eukaryotic cells.

What is meant by post-transcriptional control?

Some regulatory mechanisms that stop protein synthesis after transcription has occurred. This post-transcriptional step can also be regulated to control gene expression in the cell. If the RNA is not processed, shuttled, or translated, then no protein will be synthesized.

Define spontaneous mutation. What causes spontaneous mutation/induced mutation?

Spontaneous mutations: Occur in the absence of a mutagen. Chemicals & radiation can lead to this.

How can one amplify DNA fragments in laboratory?

The polymerase chain reaction: the process of increasing small quantities (amplifying) of DNA for analysis. Reverse-transcription PCR uses mRNA as template.

Therapeutic applications of rDNA technology.

Therapeutic applications: human enzymes and other proteins such as insulin. Subunit vaccines: made from pathogen proteins in genetically modified yeasts. Nonpathogenic viruses carrying genes for pathogen's antigens as DNA vaccines Gene therapy to replace defective or missing genes. Cervical cancer vaccine Hepatitis B vaccine Human Growth Hormone (corrects growth deficiencies in children) Gene silencing (small siRNAs bind to mRNA, which is then destroyed by RNA silencing complex) and RNA interference (inserts DNA encoding siRNA into a plasmid and transferred into a cell) could provide treatments for a wide range of diseases.

How viruses may be used to treat cancer by killing cancer cells?

These tumor-destroying viruses, or oncolytic viruses, selectively infect and kill tumor cells or cause an immune response against tumor cells. Several viruses are known to selectively infect cancer cells, and these are being genetically modified to remove virulence genes and add colony-stimulating factor genes to promote white blood cells.

catabolic repression

This can override inducible operon. Catabolic repression shows that glucose is the sugar of choice.

What is the significance of Ti Plasmid

Ti plasmid: is used as a vector for genetic modification in plants.

What is tissue culture/cell culture/monolayer? What is their use? Describe ways viruses are cultivated.

Tissue culture/cell culture: is used for Subculturing where cells from an existing culture are transferred to new containers with fresh nutrient media. A lot of separate subcultures can be made from a single tissue sample. Cells from tissue like liver and is the primary cell culture for virus-Monolayer: the cells from an organism are blended so they separate and then these cells start growing on bottom of the flask and do NOT grow on top of each other and they stop growing when close to each other-Used to make host cells for viruses and be subcultured.-You always need a host to grow viruses because they require a host to replicate.

List the enzymes and their functions involved in DNA replication.

Topoisomerase and gyrase relax the DNA strands Helicase separates the strands DNA polymerase adds nucleotides to the growing DNA strand in 5-3 direction Fragments are joined together with Ligase

Briefly describe transcription and translation. What is used as a template for each process?

Transcription: is the synthesis of a complementary strand of RNA from a DNA template. Transcription begins when RNA polymerase binds to the DNA. At the end, RNA polymerase and RNA are released. Protein synthesis is called translation because it involves decoding the "language" of nucleic acids and converting it into the "language" of proteins. The overall goal of translation is to produce proteins using mRNAs as the source of biological information.

What is the role of viruses in cancer? Name the cancers caused by viruses.

Transforming infection is when the virus has capacity to alter nucleic acid of host, leading to changes in host's DNA which can lead to drastic changes in the chromosome > cancer HPV > Cervical cancer Hepatitis B and C > Liver cancer Leukemia Lymphoma

Distinguish between transforming, persistent, latent, lysogenic, and lytic viral infection.

Transforming viral infection: virus causes CANCER, virus gets into host genes and DNA (most dangerous) and puts its DNA fragment with host DNA to make replication of viral proteins and alters the control region for cell division (have a limit of how many times they divide like kidney cells) causing UNCONTROLLABLE CELL DIVISION as a result of viral infection. (Messes up oncogene: genes controlling cell division)Ex: Human papillomaviruses (HPV) cause cervical cancer; Hepatitis B and C (HBV and HCV) cause liver cancer-Persistent viral infection: low key and virus doesn't kill the host right away and produces a small amount of new viruses (1 type is Chronic)-Chronic viral infection: DIFFERENT FROM CANCER, virus works very slow and few viruses are made and few host cells are dead (won't kill you but lingers for a long time and feel sick since hosts are dieing slowly and some viruses being produced at low level) (Ex: Hepatitis B virus (HBV) causes chronic liver disease (Drug users using dirty blood contact= I SEARCHED IT ONLINE AND HBV AND HCV CAUSES WERE FLIPPED????); Hepatitis C virus (HCV) causes chronic liver disease (Sex transmitted by body fluids and blood); Rubella virus-Latent viral infection: virus becomes part of the host but does NOT kill the host and no virus is produced, the virus goes into hiding into nerve cells and becomes latent (Ex: Varicella-zoster virus causes Chicken pox: can come back as shingles if don't have vaccine; Herpes virus (HSV) causes Herpes for rest of life will have cold sores even if not showing at the moment, it is FOREVER; Epstein-Barr Virus causes Mononucleosis a.k.a Mono; Cytomegalovirus causes subclinical thing)(few capable of doing this) (when situation turns favorable virus can go use typical lyses process and reproduce); in latency you look healthy but when virus reactivates it can produce in lyses process -- is lysogenic cycle-Lysogenic viral infection: a stable, long-term relationship btwn the phage and it's host and virus's nucleic acid becomes a part of host's nucleic acid. (viral DNA in bacteria chromosome prophage); (temperate phage and bacteria lysogen); phage (virus) adsorbed to bacteria and penetrates it, and then virus DNA inserts itself into bacteria chromosomes (prophage) and replicates with DNA replication and goes through binary fission with bacteria cell and now there are 2 virus DNAs made and it uses the host to replicate and are dormant (But if outside stimulation prophage can become active and start lytic cycle this is called induction)-Lytic viral infection: enzyme lysozyme breaks down cell wall and viruses escape and the bacteria host cell is lysed (destroyed and bursts) and now these viruses (phages) can go infect more bacteria

State the difference between and give examples of vertical and horizontal gene transfer.

Vertical gene transfer: transfer of genes from an organism to its offspring (parent to two daughter cells) Horizontal gene transfer: transfer of genes between cells of same generation. (Sideways from one bacteria to another) -Transformation (picking up DNA from the environment), transduction (virus pass DNA from one bacteria to another), conjugation (plasmids transferred from one bacterium to another via direct cell to cell contact via sex pili)

Cells prefer glucose over lactose. How are genes expressed to facilitate this preference.

When there is lactose and glucose together, the lactose operon is off. Glucose is used up and then during lag time you turn on lactose operon.

What is synthetic dna.

builds genes using a DNA synthesis.

Nanotechnology

the branch of engineering that deals with things smaller than 100 nanometers (especially with the manipulation of individual molecules)