Microbiology Exam 2 study guide
Why aren't metals used more often as antimicrobial chemicals?
its too caustic: able to burn or corrode organic tissue by chemical action.
What is meant by gene transfer and genetic recombination?
refers to the exchange of genes between two DNA molecules to form new combinations of genes in a chromosome
Differentiate between vertical and horizontal (lateral) gene transfer.
-Vertical transfer genes are passed from an organism to offspring. (Plants and animals) -Horizontal transfer genes are passed to offspring AND microbes of same generation (bacteria)
General agents usually damage microbes by one of three general mechanisms. Describe these three mechanisms of action.
1) Alteration of membrane permeability 2) Damage to proteins (structural and enzymes) 3) Damage to nucleic acids
Applications of CRISPR technology, specifically CRISPR-Cas9 1) Key steps being taken for Sickle Cell Anemia cure by researchers and physicians at Berkeley and San Francisco 2) Overview of disease 3) How CRISPR-Cas 9 is being used to exchange mutated gene with healthy gene
1) CRISPR used to correct mutation in hematopoetic stem cells of sickle cell patients 2) Overview of disease - • Single base change: glutamic acid valine. Hemoglobin binding portion of RBC's affected Making it difficult to carry O2 to lungs and rest of the body Rbcs change shape and clog capillaries so oxygen isn't delivered to tissues » can cause agacks of sudden, severe pain and organ damage. 3) By cutting DNA at site of abnormality and adhering appropriate DNA CRISPR-Cas9 is being used to correct mutation in the hematopoietic stem cells of a sickle cell patient - Promising early results but much work ahead to refine procedure - Goal is to initiate early-phase clinical trial within five years
Describe the 3 basic steps of a single PCR cycle and what is the function of each step? For example, why is the DNA sample heated to 94°C in the first step?
1) DNA incubated at 94 degrees C in order to separate the strands 2) Incubate @ 60 degrees C to allow primers to attach to single stranded DNA 3) Incubate at 72 degrees C for one min: DNA polymerase copies target DNA at this temp
Why is it better to use H2O2 on inanimate objects than on living tissue?
1) Its too caustic for living tissue. It is quickly broken down to water and gaseous oxygen by the action of the enzyme catalase which is present in human cells. 2) On inanimate objects, the normally protective enzymes of anerobic bacteria and facultative anaerobes are overwhelmed by high concetrations of peroxide. Because of these factors and its rapid degradation into harmless water and oxygen, the food industry uses it for aseptic packaging
How is H2O2 used in the hospital setting to reduce the spread of "superbugs"?
1) Sporicidal at high concentrations 2) Integrating heated, gaseous hydrogen peroxide to sterilize atmosphere and surfaces in hospital rooms using BIOQUELL. The rapid degradation of it is water and oxygen so that's good too
hybridization
1) heat to separate strands 2) Combine single strands of DNA 3) Cool to allow renaturation of double stranded DNA 4) Determine degree of hybridization Complete hybridization: Organisms identical Partial hybridization: organisms similar No hybridization: organisms unrelated
Explain the relative resistance among major microbial groups and non-living entities as discussed in class. Consider: Prions
1)Infections proteins that are the cause of neurological diseases like mad cow disease. Had to incinerate cow carcasses 2) Needs specialized autoclaving with sodium hydroxide and 134C and perhaps adding protease enzymes
Describe the process by which recombinant DNA is made and incorporated into a system to produce the product (the "product" can be either DNA or protein)
1)Vector (plasmid or virus; we will use a plasmid in our discussions for a vector) = Transports foreign DNA into a cell. Must be self replicating. Example is E. Coli being genetically modified to produce HGH.
Lysogeny 2. Specialized Lysogeny • What is it?
1. Phage is called temperate when it is in the host's DNA 2. Specialized Lysogeny • What is it? in another type of transduction, called specialized transduction, only certain bacterial genes are transferred. (in one type of specialized transduction ,the phage codes for certain toxins produced by their bacterial hosts) 3. Sometimes lysogeny results in Phage conversion (aka Lysogenic conversion): phage contains genes that make its host bacterial cell display a new trait
Donor DNA or PCR product:
Amplified quantity of DNA large enough to be analyzed Donor DNA can be a PCR product meaning that you can amplify the DNA you want to insert into the vector via PCR. You then proceed by cutting both the plasmid and the donor DNA (with gene of interest) with the SAME restriction enzyme; allowing the sticky ends of both to join together to form a recombinant plasmid (vector).
What is the fundamental difference between antimicrobial agents (chemical and physical) and antimicrobial chemotherapy (antibiotics)?
Antimicrobial Agents: chemical and physical antimicrobial agents control microbial growth in environments such as restaurants, hospitals, labs or other environments Antimicrobial chemotherapy (antibiotics): since antimicrobial agents are not useful for ingestion or injection to treat disease, antibiotics come into play.
Define: Antisepsis
Antisepsis: destruction of vegetative pathogens on living tissue]
(Antimicrobial) Damage to proteins (structural and enzymes)
Bacteria are sometimes thought of as "little bags of enzymes" which are vital for cellular activities Proteins are functional due to their three-dimensional shape, attacking the different bonds renders them dysfunctional hydrogen bonds in protein = susceptible to heat or certain chemicals
Chemical mutagen: Base analog = 5-bromouracil
Base analog: Similar to normal nitrogenous bases but slightly altered base pairing properties. They alter DNA resulting in base-pair substitutions causing a point mutation. mechanism of action: De-Aminates - causes incorrect pairing Outcome: What was supposed to be an A-T pairing turns into a C-G pairing when exposed to 5-bromouracil Type of mutation (point mutation)
Define: Biocide:
Biocide: aka germicide: treatments that cause the outright death of microbes
How is DNA fingerprinting used in forensic microbiology?
By using DNA fingerprint to track sources of attackers For several years, microbiologists have used RFLP's (restriction fragment length polymorphism) in a method of identification known as DNA fingerprinting to identify bacterial or viral pathogens. in forensic microbiology, developed because hospital, food manufactures and individuals can be sued in court and because microorganisms can be used as weapons. in the 1990's DNA fingerprints of HIV were used to obtain rape conviction. In 2001 anthrax attacks in the united states, DNA fingerprints of bacillus anthraces were used to track the source and then the alleged attacker
Sticky ends
Can be adhered to other segments of DNA unlike blunt ends some enzymes cut both strands of DNA in the same place, producing blunt ends, and others make staggered cuts in the two strands, buts that are not directly opposite each other these staggered ends or sticky ends are most useful in rDNA because they can be used to join two different pieces of DNA that were cut by the same restriction enzyme
Chemical mutagen: deaminating agents = Nitrous acid
Cause conversion of adenine to a form that pairs with cytosine instead of thymine. Results in a base-pair substitution Mechanism of action: De-Aminates - causes incorrect pairing Outcome: Point mutation
Define "Recombinant DNA"
DNA that has been formed artificially by combining constituents from different organisms. DNA in which one or more segments of DNA have been inserted, either naturally or by laboratory maipulation
Define: Degerming:
Degerming: removal of microbes from a limited area such as the skin around an injection site
Briefly describe why the following physical techniques have antimicrobial properties: desiccation
Desiccation: disruption of metabolism: involves removing water from microbes, bacteriostatic. Drypacks in food to remove excess water
Define: Disinfection
Disinfection: control directed at destroying harmful microorganisms
Explain the possible outcomes of performing a recombinant experiment using the same restriction enzyme on plasmid and Donor DNA as discussed in class.
Donor DNA: If the donor DNA has many sites that are targeted by a given restriction enzyme, then many fragments of donor DNA with sticky ends could be created - one of the fragments may contain the desired gene - other fragments aren't useful - any sticky end fragment of donor DNA could insert into plasmid Plasmid: Could reform without any donor DNA - Sticky ends "find themselves"
What obstacle had to be overcome with respect to the DNA polymerase used in PCR?
Finding the proper temperature and conditions for DNA polymerase to work use of an automated thermal cycler is made possible by the used of DNA polymerase taken from a thermophilic bacterium such as thermos aquaticus, the enzyme from such organisms can survive the heating phase without being destroyed.
What is the Origin of restriction enzymes
First isolated in 1970, restriction enzymes in nature had actually been observed earlier, when certain bacteriophages were found to have restricted host range. If these phased were used to infect bacteria other than their usual hosts, restrict enzymes in the new host destroyed almost all the phage DNA Restriction enzymes protect a bacterial cell by hydrolyzing phage DNA.
Describe the mechanism of action for H2O2 as discussed in class.
Free ions of OH break the disulfide covalent bonds. The oxidation of the various molecules that constitute microorganisms will lead to significant disruptions in structure/function and the loss of viability or infectivity.
Briefly describe why the following physical techniques have antimicrobial properties: freezing
Freezing: decreased chemical reactions and possible protein changes: bacteriostatic effect
Describe differences between eukaryotic and prokaryotic gene expression with respect to arrangement of genes involved in a metabolic pathway and how gene expression is controlled
Gene Expression: Prokaryotes: DS; single, circular; not in a nucleus Eukaryotes: DS; linear; arranged in many chromosomes in a nucleus # Alleles (copies of a gene) in chromosomal DNA Prokaryotes: 1 (Haploid) Eukaryotes: 2 (Diploid) Non-chromosomal DNA Prokaryotes: DS; Found on plasmid - small circular Eukaryotes: DS; found in mitochondria and chloroplasts; some have plasmids Is Transcription/Translation simultaneous? Prokaryotes: Yes can be simultaneously transcribing and translating Eukaryotes: Not simultaneous Arrangement of genes involved in same metabolic pathway Prokaryotes: Genes are next to each other; all controlled by one "master switch"; called an Operon Eukaryotes: Not clustered together; often on different chromosomes (i.e. spread out throughout nucleus)
Review gene expression
Gene expression is the process by which genetic instructions are used to synthesize gene products. These products are usually proteins, which go on to perform essential functions as enzymes, hormones and receptors, for example. Genes that do not code for proteins such as ribosomal RNA or transfer RNA code for functional RNA products
What does the Ames test reveal about a chemical?
If it is carcinogenic The Ames test is a widely employed method that uses bacteria to test whether a given chemical can cause mutations in the DNA of the test organism. More formally, it is a biological assay to assess the mutagenic potential of chemical compounds.
Briefly describe why the following physical techniques have antimicrobial properties: ionizing rays.
Ionizing rays: destruction of DNA by forming ions that could shatter DNA backbones, not widespread in routine sterilization Ionization of water creates highly reactive hydroxyl ions that react with organic cellular components like DNA and damaging them
Why does the plasmid vector have to carry an antibiotic resistance gene?
In order to effectively transform the donor into an antibiotic resistant organism the plasmid vector used contains a gene encoding resistance to the antibiotic ampicillin. The host bacteria won't be able to grow on the test medium containing ampicillin, unless the vector has transferred the ampicillin resistance gene.
Explain the concept of an operon.
In the control region of the lac operon there are two relatively short segments of DNA. One, the promotor is the segment where RNA polymerase initiates transcription. The other is the operator which is like a traffic light, that acts as the go or stop signal for transcription of structural genes a set of operator and promotor sites and age structural genes they control define an OPERON.
Between spontaneous and induced mutation, which type has a higher mutation rate?
Induced mutation a mutagen usually increases the spontaneous rate of mutation, which is about one in 10x6 in replicated genes, by a factor of 10 to 1000 times. in other words, in the presence of a mutagen, the normal rate of 10x-6 per genes replicated becomes 10x-5 to 10x-3
Distinguish between an inducible operon and a repressible operon.
Inducible operons are transcribed only when they are activated. -Inducible operons are off until presence of something triggers them to turn on. Repressible operons are always transcribed unless they are deactivated by a repressor. -In repressible operons, structural genes are transcribed until they are turned OFF. Ie: tryptophan
How does genetic recombination contribute to genetic diversity?
It allows for new combinations of genes on a chromosome like mutation, genetic recombination contributed to a populations genetic diversity. in highly evolved organisms such as present fay microbes, the recombination is more likely to beneficial than mutation because recombination will less likely destroy a genes function and may bring together combinations of genes that enable the organism to carry out a valuable new function
What does recombinant mean?
It means of or resulting from new combinations of genetic material. Once transferred, part if the donors DNA is usually incorporated into the recipients DNA, the remainder is degraded by cellular enzymes. the recipient cell that incorporated donor DNA into its own DNA is called recombinant
Is a mutagen necessarily a carcinogen?
No As many mutations can cause cancer, mutagens are therefore also likely to be carcinogens, although not always necessarily so.
(Antimicrobial) Alteration of membrane permeability
Membrane actively regulates the passage of nutrients into the cell and elimination of wastes. Damage to the membrane causes cellular contents to leak into the surrounding medium and interferes with the growth of the cell
Do you find operons in prokaryotes or eukaryotes?
No, only in prokaryotes
Briefly describe why the following physical techniques have antimicrobial properties: heat (moist and dry),
Moist Heat: protein denaturation: kills vegetative bacterial and fungal pathogens and almost all viruses within 10 min, less effective on endospores AUTOCLAVE: 15 psi (pounds per square inch) at 121C for 15 mins Dry Heat: direct flaming, burns contaminants into ashes: effective sterilization
Ophthalmia neonatorum:
Neonatal conjunctivitis, also known as ophthalmia neonatorum, is a form of conjunctivitis and a type of neonatal infection contracted by newborns during delivery. The baby's eyes are contaminated during passage through the birth canal from a mother infected with either Neisseria gonorrhoeae or Chlamydia trachomatis.
Looking back at the new technologies used in the clinical lab (from L3), how do "PCR in a box" and "NAT" provide identifications of microbes?
PCR- Individual cartridges contain all the ingredients to run PCR including primers that delineate a gene specific to the pathogen. NAT- The cartridge contains numerous probes that are specific to each pathogen
Describe DNA repair enzymes: photolyase and nucleotide excision repair.
Photolyase uses visible light energy to separate thymine dimers Nucelotide excision repair- enzymes cut out incorrect base and fill gap with new DNA
Transformation
Process by which plasmid is inserted into a cell during which cells can take up DNA from the surrounding environment.
Describe differences between eukaryotic and prokaryotic gene expression with respect to a. arrangement of genes involved in a metabolic pathway and b. how gene expression is
Prokaryotes: controlled bacteria grows in competitive rapid conditions, Simple needs and single celled, don't have specific gene expression, have to respond quickly to environmental changes, organized in operons and transcribed by a single Mrna, polysystronic Mrna, transcription and translation can occur at same time Eukaryotes: have other regulatory needs so they have a different set of regulatory controls, NOT regulated by operons, each structural gene has its own promoter and is transcribed independently, monocistronic, chromatin structure affects gene expression, have more transcriptional activators as well as gene repressors, transcription/translation physically separated
What is the main goal of the polymerase chain reaction (PCR)?
Purpose: Amplify (Increase) small amounts of DNA : that is, increased quantities that are large enough for analysis PCR is a common tool used in medical and biological research labs. It is used in the early stages of processing DNA for sequencing, for detecting the presence or absence of a gene to help identify pathogens during infection, and when generating forensic DNA profiles from tiny samples of DNA.
What bacterial enzymes are used in genetic engineering?
Restriction enymes and CaS
Explain how restriction enzymes reflect an "innate" immunity for bacteria and CRISPR reflects an "adaptive" immunity in bacteria.
Restrictive enzymes occur naturally (you are born with innate immunity), whereas CRISPR is a synthetic process (that adapts to environmental changes as part of evolution)
Briefly describe why the following physical techniques have antimicrobial properties: high concentration of salt or sugar
Salt and sugar form hypertonic solution in which water concentration is lower but water always goes higher concentration to lower concentration(osmosis), so, water from microbes goes out due to osmosis. and due to dehydration microbes die.
Describe a medical application of AgNO3 (silver nitrate) in a neonatal unit.
Silver nitrate was smeared on the eyes of newborn infants to prevent transmitted gonnhorea and chlamydia from attacking their eyes. Ophthalmia neonatorum: This helped reduce congenital blindness, but was replaced with an antibiotic smear because the silver nitrate became too caustic.
What does turning on a gene mean? What does turning off a gene mean? Is control over gene expression exerted on the transcription or translation level?
Some genes are on until they are turned off (always functioning until signaled not to). Others are off until stimulated to turn on. This is a means of metabolic efficiency.
what are restriction enzymes?
Special class of DNA cutting enzymes that exist in many bacteria. Protect bacteria by hydrolyzing phage DNA
Differentiate between a spontaneous mutation and an induced mutation.
Spontaneous mutations occur spontaneously because of occasional change in base sequence of DNA. Induced mutation happens after exposure to a mutagen. Mutations may be induced by exposure to ultraviolet rays and alpha, beta, gamma, and X radiation, by extreme changes in temperature, and by certain mutagenic chemicals such as nitrous acid, nitrogen mustard, and chemical substitutes for portions of the nucleotide subunits of genes.
Explain the relative resistance among major microbial groups and non-living entities as discussed in class. Consider: Presence of spores
Spores are generally resistant to biocides
Define: Sterilization
Sterilization: removal and destruction of all living microorganisms (usually heating)
Briefly explain why we use antiseptics and not sterilizers on living tissues prior to invasive medical procedures.
Sterilizers would be too harsh on living tissues, you would damage the cells of the tissues that you're trying to protect.
how does CRISPER-Cas9 work?
The CRISPR-Cas9 system consists of two key molecules that introduce a change (mutation) into the DNA. These are: an enzyme called Cas9. This acts as a pair of 'molecular scissors' that can cut the two strands of DNA at a specific location in the genome so that bits of DNA can then be added or removed. a piece of RNA called guide RNA (gRNA). This consists of a small piece of pre-designed RNA sequence (about 20 bases long) located within a longer RNA scaffold. The scaffold part binds to DNA and the pre-designed sequence 'guides' Cas9 to the right part of the genome. This makes sure that the Cas9 enzyme cuts at the right point in the genome
Name the three mechanisms of bacterial horizontal gene transfer. (AKA lateral gene transfer) and be able to differentiate among the three mechanisms of horizontal gene transfer.
Transformation: Genes are transferred from one bateria to another as naked DNA. Conjugation: Genetic material transferred via plasmid. Requires cell to cell contact. Transduction: Bacterial DNA transferred from a donor cell to recipient cell inside a virus that inserts bacteria.
(Antimicrobial) Damage to nucleic acids
The nucleic acids DNA and RNA are the carriers of the cell's genetic information. Damage to these nucleic acids by heat, radiation, or chemicals is frequently lethal to the cell. the cell can no longer replicate, nor can it carry out normal metabolic functions such as the synthesis of enzymes
How do RFLPs relate to DNA fingerprinting-They allow for any person's DNA to be tested for the presence of a mutated gene
The sequence of a mutated gene can be used as a diagnostic tool in hybridization technique called southern blotting. In this technique, subject DNA is digested with a restriction enzyme, yielding thousands of fragments of various sizes. The fragments are called RFLP's the different fragments are then separated by gel electrophoresis. The fragments are put in a well at one end of a layer of agarose gel. then electrical current is passed through the gel. while charge is applied, the different sized RFLPs migrate through the gel at different rates. the RFLP's are transferred onto a filter by blotting and are exposed to a labeled probe made from the glued gene of interest. the probe with hybridize to this mutant gene but not the normal gene. fragments to which the probe binds are identified by colored dye with this method, any persons gene can be tested for the presence of a mutated gene.
Is control over gene expression exerted on the transcription or translation level?
The term 'gene expression' is sometimes used to refer to the transcription phase alone. Genes are exerted at the level of transcription.
How is gene expression controlled in prokaryotic cells vs eukaryotic cells?
Therefore, in prokaryotic cells, the control of gene expression is mostly at the transcriptional level. In eukaryotes, The regulation of gene expression can occur at all stages of the process. Regulation may occur when the DNA is uncoiled and loosened from nucleosomes to bind transcription factors (epigenetics), when the RNA is transcribed (transcriptional level), when the RNA is processed and exported to the cytoplasm after it is transcribed (post-transcriptional level), when the RNA is translated into protein (translational level), or after the protein has been made (post-translational level).
Mutagen: Physical: UV Radiation
UV radiation: Formation of harmful covalent bonds between pyrimdine bases resulting in adjacent thymines called thymine dimers. This causes cell damage or death. Outcome: ranges: from point mutation to frameshift mutation
Briefly describe why the following physical techniques have antimicrobial properties: UV rays
UV rays: damage to DNA, creates the Thymine-Thymine bridge bulge, DIMERIZATION
What steps must be taken to know for sure that your experiment was successful?
When gene of interest is interested into vector -Vector must contain antibiotic resistant gene -Direct selection determines if plasmid got into host cell -Colony hybridization detects gene of interest in a colony only bacteria that picked up the plasmid will grow, because they are now ampicillin resistant.
Mutagen: Physical: X-ray
X-ray radiation: production of free radicals causng DNA errors or breaking of covalent bonds in backbone of DNA which results in physical breakdowns of chromosomes. Mechanism of action : Ions formed when exposed to X rays Outcome: Varies: from base pairing mistakes to breakage of DNA backbone
Explain the relative resistance among major microbial groups and non-living entities as discussed in class. Consider: Enveloped versus non-enveloped viruses
a) Antimicrobials that are lipid-soluble are more likely to be effective against enveloped viruses b) Nonenveloped viruses are more resistant because they have a protein coat
a) What is transduction? b) How can a bacterial virus (phage) donate bacterial genes to a host bacterium? c) Lytic cycle d) Lysogen Phage conversion e)How does phage conversion contribute to an organism's pathogenicity? f) Give two examples of organisms that have become pathogenic due to phage conversion.
a) Bacterial DNA transferred from a donor cell to recipient cell inside a virus that infects bacteria, called a bacteriophage or a phage. b) 1) phage infects bacterial cell 2) phage DNA and proteins are made and the bacterial chromosome is broken into pieces 3) during phage assembly pieces of bacterial DNA are packaged in phage capsid. then the donor cell lyses and releases phage particles containing bacterial DNA 4) phage carrying bacterial DNA infects a new host cell, the recipient cell 5) recombination can occur, producing a recombinant cell with genotype different from both the donor and recipient cells c) Lysis of the cell and release of phage particles. Leads to production of new phage particles which are released by lysis of the host. d) A lysogen or lysogenic bacterium is a bacterial cell in which a phage exists as DNA in its dormant state Phage conversion = Induces a change in phenotype of infected bacteria that is not part of usual cycle. e) It potentially makes them more pathogenic f) Corynebacterium diptheriae and Clostridium botulinum
Explain the relative resistance among major microbial groups and non-living entities as discussed in class. Consider: a) cell wall composition of microbes
a) Biocides tend to be more effective against gram+ bacteria than gram -. 1)Lipopolysaccharide layer of gram- is more resistant and their porins 2)Mycobacteria have a waxy, lipid-rich component
a) What is CRISPER b) How did it originate? c) How does CRISPER-Cas work?
a) Clustered regularly interspaced short palindromic repeats b) Originated in form of immunity in bacteria c) "spacer" sequences are transcribed into short RNA sequences capable of guiding the system to matching sequences of DNA. When target DNA is found, Cas9, an enzyme produced by CRISPR system, finds to DNA and cuts it, shutting the targeted gene off
a) What is filter sterilization? b) When does one need to use filter sterilization?
a) Filtration is the passage of liquid or gas through a screenlike material with pores small enough to retain microorganisms. Used for sterilizing heat-sensitive materials like culture media, enzymes, vaccines, and antibiotic solutions. Two chamber apparatus with air pressure pulling the liquid from the top through the bottom, change the filtration size to make sure the small bacteria/viruses can't pass through. b) for heat-sensitive materials (heat-labile)
Heat as a physical method of sterilization can be moist or dry a) What are examples as discussed in class? b) How does an autoclave work? c) How does incineration work?
a) Flaming inoculation loop vs autoclaving materials b) Pressure allows you to achieve higher temperature and the steam can kill microbes by degradation c) a similar principle to inoculation loop sterilization, incineration is an effective way to sterilize and dispose of contaminated paper cups, bags and dressings
a) What is transformation? b) Describe Fredrick Griffith's experiments that first demonstrated "transformation". c) What is competency? d)Are all organisms naturally competent? e)How do you make an organism become competent? f) What type of DNA can be "taken up" by a bacterium? (i.e. chromosomal or plasmid)
a) Genes are transferred from one bateria to another as naked DNA. b) tested whether injections of heat killed bacteria of encapsulated strain could vaccinate mice against pneumonia. Injections of living encapsulated bacteria killed the mouse, but injections of live non encapsulated or dead encapsulated bacteria did not kill the mouse. however, when the dead encapsulated bacteria were mixed withe the live non encapsulated bacteria, many of the mice died. c) when a recipient cell is in a physiological state in which it can take up donor DNA, it is said to be competent. d) No e) competence results from alterations in the cell wall that make it permeable to large DNA molecules f) Chromosomal DNA
a) What is conjugation? b)What are F+ and Hfr? c) Differentiate between the F+→F- & Hfr → F- methods of conjugation . d) What can be transferred (e.g. plasmid and/or chromosomal DNA)?
a) Genetic material transferred from one bacterium to another is known as conjunction, its mediated by one kind of plasmid, a circular piece of DNA that replicated independently from the cells chromosome b) F factor = (fertility factor) Hfr = high frequency of recombination c) donors caring F+ transfer plasmid to recipient F- cells which become F+ cells as a result In some cells carrying F factors, the factor integrates into the chromosome, converting the F+ cell to an Hfr cell. When conjunction occurs between and Hfr cell and an F- cell, the Hfr cell's chromosome (with its integrated F factor) replicated and a parental strand of the chromosome is transferred to the recipient cell. therefore, by conjunction withe an Hfr cell and an F- cell may acquire new versions of chromosomal genes, however it remains an F- cell because it did not receive a complete F factor during conjugation d) Both chromosomal DNA and Plasmid for Hfr and just plasmid for F+
Describe the mechanism of action for detergents (e.g. soap). Are detergents disinfectants, sterilizers, or degerming agents?
a) Soaps and detergents are amphipathic (positive charge and have a hydrophobic region) and disrupt the lipid bilayer. b) Surfactant: surface active agent decreases surface tension among molecules of a liquid c) DEGERMING AGENTS: soap mechanically breaks it down and lifts debris
a) What does "oligodynamic action" mean? b) Describe the mechanism of action for heavy metals as described in class.
a) VERY LITTLE (very little amounts in order to be effective) Ability of heavy metals, in small amounts, to exert antimicrobial activity usually silver, copper and mercury b) Extremely small amounts of metal diffuse from the source and inhibit the growth of bacteria because of the metal ions are combining with sulfhydryl groups on cellular proteins Protein denaturation: • Disrupts disulfide bonds • Disrupts salt bridges
Chemical mutagen: Acridine dyes
acridine dyes: Intercalate into DNA causing frameshift mutation. Mechanism of action: Intercalating agents - (wedges itself into double helix and causes frameshift mutation) Outcome: Point mutation
Explain the mechanisms of action for alcohol as described in class. (Alcohol can both denature proteins and disrupt membranes.)
alcohol usually denatures protein, but it can also disrupt membranes and dissolve many lipids, including the lipid component of enveloped viruses they have the benefit of acting and then evaporating rapidly and leaving no residue
Terms: colony hybridization Probes
colony hybridization is a common method of identifying cells that carry a specific cloned gene. DNA probes, short segments of single stranded DNA that are complimentary to the desired gene, are synthesized. If the DNA probe finds a match, it will adhere to the target gene. The DNA probe is labeled with an enzyme or fluorescent dye so its presence can be detected.
Cure of a disease in pigs called Porcine Reproductive and Respiratory Syndrome (PRRS); a viral disease How was CRISPR-Cas9 used to protect pigs from infection by virus?
edited OUT the receptor for the virus responsible for causing this disease in pigs • Results showed that pigs that were "edited" were not infected with the virus when exposed whereas "unedited" pigs were infected and developed the disease.
What does the acronym RFLP mean?
restriction fragment length polymorphous
Which type is the lac operon?
the combination of the 3 lac structural genes and the adjoining control regions is called the lac operon In lac operon, in absence of lactose, repressor binds to operator site, preventing transcription. In presence of lactose, binds to a metabolite of lactose instead and enzyme is transcribed.
What concentration of alcohol is needed to be effective?
the recommended optimum concentration of ethanol is 70%, but concentrations between 60-95% seem to kill as well. pure ethanol is less effective than aqueous solutions because denaturation requires water.
Define: Sanitization: treatment intended to lower microbial counts on eating and drinking utensils to safe public health levels food industry
treatment intended to lower microbial counts on eating and drinking utensils to safe public health levels food industry
Explain how a DNA fingerprint experiment is done
via PCR microarrays that can screen a sample for multiple pathogens at once. Up to 22 primers from different microorganisms can be used. A suspect microorganism is indentified if DNA is copied from one of the primers.
