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AnswersSampleExam1_1 - Sample Questions and Answers...

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Unformatted text preview: Sample Questions and Answers For Exam 1 1. You have isolated a archeal species that lives in a hydrothermal vent at 90°C. You have determined that this archea has an S ­layer. You find that you can remove the S ­layer by treating the cells with a protease. To your surprise, you find that the S ­layer is not required for survival in low osmolarity (i.e. low solute concentration) environments. You hypothesize that the S ­layer is critical for these archeal cells to survive at the temperatures at which it grows, 90°C. Outline an experiment to test this hypothesis. Be sure to include a control in your experiment, and list the results you would expect if your hypothesis is correct. Place your cells in a buffer or growth medium. Incubate the cells at 90°C. Collect samples at various time intervals. (not a critical component, an endpoint experiment would be sufficient) Make dilutions of the samples and plate to measure CFU/ml. Control group: cells with the S layer Experimental group: cells with out the S layer Expected results: The cells with the S layer will yield a constant level of CFU/ml versus time. The cells with out the S layer will show a decrease in CFU/ml versus time. 2. You notice that there is a rod ­shaped bacterium present in the blood of the 10 sheep with a debilitating motor disease. This bacterium is not found in the blood of 10 sheep that do not have the motor disease. You name this bacterium S. motorus, and you suspect that it is responsible for causing the motor disease. You streak purify S. motorus, and inject a pure culture of this organism into 10 sheep that appear healthy. All 10 of the injected sheep develop the disease and you are able to re ­isolate S. motorus from 9 of the injected sheep. You conclude that S. motorus is the causative agent of the motor disease. You then find that another laboratory has data to support the motor disease being caused by a virus, not S. motorus. Indicate what is a possible flaw in your experimental technique that led you to incorrectly conclude that S. motorus is the causative agent of the motor disease. You should have included a group of control sheep that were not injected (or injected with just buffer). This would have indicated whether the group of sheep was already infected (with another disease causing agent) and the disease symptoms had not yet manifested themselves. 3. You want to isolate a bacterium that uses cellulose as a carbon and energy source. You have a soil sample taken from a field with decaying plants. You hypothesize that your desired bacterium will be relatively rare in this sample. You design a defined growth medium whose only added source of carbon is cellulose. a) Indicate whether you would use a liquid or solid form of the growth medium to enrich for cellulose ­degrading bacteria and Why? LIQUID MEDIUM  I would choose the liquid medium because the cellulose degrading bacterium should grow faster than the other bacteria. I would not choose the solid medium because the cellulose degrading bacteria are so rare that plating a small portion of the environmental sample on the plate would not result in growth of any of the desired bacteria. b) You want to know whether the prokaryotic organisms you have isolated from your enrichment are Gram ­negative bacteria, Gram ­positive bacteria or Archea. Indicate what you would do to distinguish this? A1: 16s rRNA gene sequencing A2: Characterize membrane lipids and perform Gram stain. c) You determine that all the bacteria isolated are Geobacillus stearothermophilus. You want to know whether these are all the same strain of G. stearothermophilus or whether they are different. Outline a procedure to determine the relatedness of the different G. stearothermophilus isolates. Isolate chromosomal DNA from each strain. Digest the DNA with a restriction enzyme. Run the digested DNA on a pulse field gel. (regular gel  0.5 points) Compare the pattern of DNA fragments for each strain. d) G. stearothermophilus is a Gram ­positive bacterium, that you find is able to specifically adhere to cellulose. You hypothesize that G. stearothermophilus produces a protein to that specifically binds cellulose and that is responsible for mediating binding of G. stearothermophilus cell to cellulose. In which subcellular structure/location would you expect the cellulose adhesion molecule be located? (Choose one) 1. Cytoplasm 4. Periplasm 2. Cytoplasmic membrane 5. Outer membrane 3. Peptidoglycan e) You examine your G. stearothermophilus strains by transmission electron microscopy. You observe a structure outside of the presumed peptidoglycan layer that you hypothesize is a capsule. What test would you carry out to verify whether your G. stearothermophilus strain has a capsule? What result would you observe if your hypothesis is correct? Use india ink to negatively stain a culture of G. stearothermophilus and then view it by bright field light microscopy (or with a compound light microscopy or phase contrast scope). If the cells have a capsule, there will be a large clear area around the cells in contrast to the dark background. 4. (Fill in the blank; one word answers) The function of the Gram ­negative bacterial lipoprotein is to cross ­links the outer membrane to the ______________ (peptidoglycan). Small molecules cross the outer membrane through ______________ (porins). 5. Many bacteria produce alcohols from sugar in the process of energy generation. These alcohols are toxic to cells. Alcohols can passively diffuse across a membrane, but several bacterial species have an antiporter to export alcohols. What advantage would having an alcohol ­specific antiporter provide for these bacteria versus those bacteria that simply relying on passive diffusion of alcohols across the membranes? An antiporter uses energy to drive transport of compounds. This allows the cells to achieve a lower concentration of the transported substrate, in this case, alcohols, inside the cell relative to the concentration that could be achieved through simply passive diffusion. (Acceptable alternative: higher concentration outside the cell.) In passive diffusion, the intracellular concentration will come to equilibrium with the exterior concentration. 6. You have two different bacterial species that both utilize glucose as a sole carbon and energy source. Species A uses a PTS system transporter for glucose; whereas species B uses an ABC transporter for glucose. You mix a small, but equal numbers of cells together of species A and species B and grow them together in a defined medium with glucose as the sole carbon and energy source. You incubate this mixed culture to allow growth of the cells, at the end of growth do you expect: (choose one) a) equal numbers of species A and species B cells b) greater number of species A cells c) greater number of species B cells 7. You have two different bacterial species, one of which, species A, uses a symporter to transport glucose and the second of which, species B, uses an ABC transporter for glucose. You mix equal amounts of the species A and species B cells in a defined minimal medium containing 10 µM glucose (i.e. a low concentrations) as the sole carbon source. After incubation for several hours, would you expect to find: (Choose one) a) equal numbers of species A and species B cells b) greater number of species A cells c) greater number of species B cells 8. You have found that the soil bacterium, P. fluorescens, can use the pesticide KillX as a source of Nitrogen for growth. You hypothesize that KillX is actively transported into the cell. Outline an experiment to test your hypothesis. Be sure to include a control in your experiment. Incubate cells with different concentrations of radioactive KillX. At different times, remove a sample of cells. Separate the cells from the free, radioactive KillX. Determine the amount of radioactivity associated with cells. Determine a rate by dividing the amount of radioactivity associated with the cells by time of incubation (or the change in the amount of cellular radioactivity divided by the change in time). Plot the measured rate versus the concentration of KillX at which that rate was measured. – 5 points Experimental Group: Incubate B. japonicum with different concentrations of radioactive KillX. Control Group: Incubate radioactive KillX in the absence of cells. OR Incubate cells with out radioactive KillX. – 2 points for the experimental and control groups. 9. You have isolated a new species of bacterium (K. pollutanti) from a site contaminated with trichloroethylene (TCE) and are able to grow it in pure culture. You examine K. pollutanti by electron microscopy and determine that it has flagella like those found on E. coli. You hypothesize that K. pollutanti is able to colonize TCE ­contaminated sites by chemotaxing toward TCE. Describe an experiment you would use to test your hypothesis that TCE acts as an attractant for K. pollutanti. Be sure to describe the result you expect if TCE is an attractant, as well as what you expect if it is a repellent or neither. Include a control. Acceptable answers:  Perform a capillary assay using trichloroethylene as the test compound. If trichloroethylene is an attractant, K. pollutanti should accumulate in the capillary tube. If trichloroethylene is not an attractant, K. pollutanti should accumulate away from the capillary tube (repellent), or remain evenly distributed (neutral). As a negative control, use medium alone. (Note, for this answer the assay should be drawn out, or described in greater detail.)  Other well described alternatives, e.g. plating and adding spot of attractant, will be considered. 10. Underline which of the following statements are true: If at the end of a run, an E. coli cell finds itself in a higher concentration of attractant than at the beginning of the run, the E. coli cell would always tumble and then run… a) …in the direction in which it is now pointed and for a shorter time period than the previous run. b) …in the direction in which it is now pointed and for a longer time period than the previous run. c) …in the same direction and for a longer time period than the previous run. d) …in the same direction and for a shorter time period than the previous run. 11. What is autotrophic growth? (Choose one answer.) a) growth using CH4 as the sole carbon source b) growth using CH4 as an electron donor for respiration c) growth using CH4 as an electron acceptor for respiration d) growth using CO2 as the sole carbon source e) growth using CO2 as an electron donor for respiration f) growth using CO2 as an electron acceptor for respiration 12. Nitrogen fixation and CO2 fixation are processes that lead to the conversion of inorganic molecules into biomass. Both these processes are energetically expensive, requiring a large number of ATP and reducing equivalents. List two general ways in which nitrogen fixation and CO2 fixation differ. Multiple mechanisms have evolved for CO2 fixation, but only one mechanism has evolved for nitrogen fixation. Nitrogen fixation is a pathway, and most mechanisms for CO2 fixation are cycles. 13. You inoculate a growth medium with equal numbers of an obligate heterotrophic bacterium and an obligate autotrophic bacterium. The only source of carbon or nitrogen in the growth medium is atmospheric CO2 and N2. The energy source of provided for the culture is light. At the end of the incubation period, you find that the numbers of both the heterotrophic bacterium and the autotrophic bacterium have increased. Speculate as to the carbon and nitrogen sources used by the heterotrophic bacterium for growth. The carbon and nitrogen for the heterotrophic bacterium most likely came from dead cells of the autotrophic bacterium. Protein, lipid, DNA, or polysaccharides could have provided the carbon and nitrogen for the heterotrophic bacterium. 14. You have cultures that each have two species of microorganism present in them. One culture is an enrichment of an environmental sample for microorganisms that can utilize ethanol as a sole carbon source. The second culture is an enrichment of an environmental sample for microorganisms that can utilize butyrate as a sole carbon source. (Ethanol and butyrate are both common end-products of primary fermentation pathways.) For each culture, neither species present in the culture can grow using either ethanol or butyrate as the sole carbon source. You sequence the genes for the rRNA from present in the two cultures. The ethanol enrichment culture contains species you call Species A and Species B. The butyrate enrichment culture contains a Species C and Species B. Thus, in both cultures Species B is present. Assuming that Species B serves the same function in the growth of both the cultures, speculate as to the function at Species B may be serving for these cultures. Species B is a hydrogen-consuming organism (or methanogen) that keeps the partial pressure of hydrogen low and thus allows the reactions carried out by the other species to be energetically favorable. 15. The organic carbon compound, glucose, is converted to an inorganic carbon compound under aerobic conditions and under anaerobic conditions even in the absence of any terminal electron acceptor for respiration. A. What are the inorganic carbon compounds that are the end-products of this conversion… Under aerobic conditions? CO 2 Under anaerobic conditions? CH4 OR CH4 and CO2 B. How many different organisms are required to convert glucose to the inorganic endproducts… Under aerobic conditions? One Under anaerobic conditions? Three 16. Explain why is denitrification harmful to plant growth in agricultural settings? Because denitrification converts nitrate to gaseous forms of nitrogen that enter the atmosphere and thereby deplete the soil of nitrogen. 17. Nitrification and denitrification serve a similar function for cells in allowing cell growth. What is this function? Both are respiration pathways that generate a PMF and, through that, ATP. 18. Leguminous plants can form a symbiotic relationship with nitrogen-fixing species of rhizobacteria when N2 gas is the only form or nitrogen present. Given your knowledge of nitrogenase, indicate what is one critical environmental condition that the plant provides for the rhizobacteria to fix nitrogen and why this environmental condition is critical. The plant keeps the levels of oxygen low so as not to inactivate nitrogenase. 19. Describe the process of replication of the genome of a ssRNA( ­) virus including which enzyme(s) are involved in this process. The negative strand genome is copied to a positive strand by an RNA dependent, RNA polymerase. This positive strand then serves as the template for synthesis of new negative strand copies of the genome. Transcription of the positive strand to the negative strand also uses an RNA dependent RNA polymerase. 20. You are studying a newly discovered mammalian virus. In the viral particles, a ssRNA strand exists that has terminal repeats. What do these terminal repeats suggest about the class of virus to which this new virus belongs? a) Group I: dsDNA b) Group II: ssDNA c) Group III: dsRNA d) Group IV: ssRNA (+) e) Group V: ssRNA ( ­) f) Group VI: Retrovirus g) Group VII: dsDNA pararetrovirus 21. Which type of protein(s) is likely to be expressed late in the infection cycle of a virus (choose all that apply)? a) polymerase b) capsid protein c) lytic proteins d) maturation proteins 22. You have identified a new dsDNA virus. One of the putative proteins encoded by the genome is similar to proteins that are known to have DNA nicking activity. What do you propose the type of replication to be for this genome? Rolling circle replication 23. You have identified a new ssRNA( ­) virus of mammalian cells. One of the proteins you find in the viral particles is a RNA endonuclease. You propose a new mechanism for production of multiple proteins from a single ssRNA( ­) molecule – the endonuclease acts on the ssRNA (+) strands, created from the ssRNA ( ­) genome, to generate multiple ssRNA (+) fragments each of which will code for one protein. For this mechanism of multiple protein expression to work, what other functions will this virus need? It will need a mechanism to ensure poly adenylation of the 3’ end of the ssRNA (+) fragments. It will also need either a mechanism to add 5’ caps to the ssRNA (+) fragments or each of the ssRNA (+) fragments should have a IRES element near their 5’ end. 24. The ribosome ­binding site (RBS) for the maturation protein of an ssRNA (+) virus of a bacterium folds into a complicated stem ­loop structure that makes the RBS unavailable to ribosomes. Describe a mechanism that this virus may use to ensure that a small amount of the maturation protein is produced. The stem loop structure is melted and the RBS made available when RNA polymerase copies the ssRNA (+) strand to the ssRNA ( ) strand. As RNA polymerase will only make a small number of ssRNA ( ) strands, the RBS for the maturation protein will only be rarely available. Thus, only small amount of maturation protein will be made. ...
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