Genetics - Analysis and Principles

Biology

Quiz 6 :

Genetic Linkage and Mapping in Eukaryotes

Quiz 6 :

Genetic Linkage and Mapping in Eukaryotes

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In a conjugation experiment, what is meant by the time of entry How is the time of entry determined experimentally
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Conjugation is the transfer of genetic material from one bacterium to other with the help of conjugating bridge. This bridge is also known as sex pilli. Through this channel, F plasmid is moved from donor (F + cell) into recipient cell (F - ). Mr. EW FJ conducted an experiment to map the genes on the chromosomes. As mentioned, such transfer of genetic material with the help of physical contact appears to be as mating. They found that during the mating of bacteria, by vigorous shaking, the bacteria get separated from each other and conjugation gets disrupted. During this process of shaking, no bacteria get harmed. Such disrupted mating is called as interrupted mating. They also hypothesized that characters or genes on DNA strand are linearly arranged. During conjugation, genetic material will take some time to move from F + cell to F - cell. If mating is interrupted early, it was found that only a few genes got transferred. When conjugation was disrupted after a bit long time, F plasmid which managed to enter recipient cell had more genes than beforE.They used the units of minutes to measure the distance between genes. A time taken for one gene to enter recipient cell is mentioned as time of entry. When mating was disrupted in short period of time during conjugation, it was found that only one gene got transferred. With the help of frequent interruption, time of entry of each gene was estimated.
In an experiment, the F plasmid of E. coli is allowed to move from donor cell to recipient cell during conjugation. After 16 minutes, conjugation was interrupted and found that only lacZ gene got along with plasmid. Further, the experiment was continued and bacterial mating was disrupted after 25 minutes. It was found that along with lacZ gene, galE gene also got into recipient cell. With the graph plotted, it was found that difference between (25-16 = 9 mins) the time taken to enter F - cell is actually the distance between these two genes. This also explains that time to enter recipient cell of lacZ gene is 16 mins, while of galE gene is 25 mins.

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Think about the structure and transmission of F factors and discuss how you think F factors may have originated.
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The extrachromosomal genetic material of E. coli is called as the plasmid or F plasmid (Fertility factor or sex factor). This plasmid is independent of having all the sufficient genes for replication, forming conjugating bridge, and transferring a single strand of this plasmid from one cell to other. The oriT region of the F plasmid will form a nick and with the help of protein relaxosomes, a single strand of DNA will roll out. The traA gene of the plasmid will form a sex pilus structure, which will enable bacteria to bind with the recipient cell (F - cell). The sex pilus will form a conjugating bridge through which the single-stranded DNA will move into the F - cell.
This F plasmid is usually a small covalently closed circular double-stranded DNA. The size of the plasmid is almost 2 percent (94.5 kb) of the bacterial chromosome. It can code about 94 proteins. This shows that this plasmid was once part of the bacterial chromosome. During replication (rolling circle) of the bacterial chromosome, some part of DNA having the oriT region gets detached accidently and thus fragment forms a double-stranded circular DNA with the help of replicating machinery present on the broken fragment. Hence, this fragment becomes stable to form a F plasmid. Since, this plasmid can get easily mixed with other genetic elements, with time plasmid acquires qualities, such as antibiotic resistance, temperature resistance, etc.

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What is the role of sex pili during conjugation
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The genetic material is transferred from one bacterium to another with the help of a physical contact which is known as conjugation. During this mechanism, the donor cell (the bacterium, which delivers DNA) forms a structure, which helps in attaching physically with another bacterium (recipient cell). This structure is called sex pili (singular: pilus). The donor cell is also known as the F + cell. With the help of sex pili, the F + cell will form a conjugating bridge with the F - cell. Through this F plasmid will be transported from the F + cell into the F - cell. The sex pilus is formed of the pilin protein, which is encoded by a gene present on the F plasmid ( traA gene).

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What is the difference between an F + and an Hfr strain Which type of strain do you expect to transfer many bacterial genes to recipient cells
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Each species of bacteria has its own distinctive cell surface. The characteristics of the cell surface play an important role in processes such as conjugation and transduction. For example, certain strains of E. coli have pili on their cell surface. These pili enable E. coli to mate with other E. coli , and the pili also enable certain bacteriophages (such as M13) to bind to the surface of E. coli and gain entry into the cytoplasm. With these ideas in mind, explain which forms of genetic transfer (i.e., conjugation, transduction, and transformation) are more likely to occur between different species of bacteria. Discuss some of the potential consequences of interspecies genetic transfer.
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The terms conjugation, transduction, and transformation are used to describe three different natural forms of genetic transfer between bacterial cells. Briefly discuss the similarities and differences among these processes.
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Explain how a U -tube apparatus can distinguish between genetic transfer involving conjugation and genetic transfer involving transduction. Do you think a U -tube could be used to distinguish between transduction and transformation
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Complementation occurs when two defective alleles in two different genes are found within the same organism and produce a normal phenotype. What other examples of complementation have we encountered in previous chapters of this textbook
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What is the role of the origin of transfer during F + - and Hfr-mediated conjugation What is the significance of the direction of transfer in Hfr-mediated conjugation
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Acridine orange is a chemical that inhibits the replication of F factor DNA but does not affect the replication of chromosomal DNA, even if the chromosomal DNA contains an Hfr. Let's suppose that you have an E. coli strain that is unable to metabolize lactose and has an F factor that carries a streptomycin-resistant gene. You also have an F strain of E. coli that is sensitive to streptomycin and has the genes that allow the bacterium to metabolize lactose. This second strain can grow on a lactose-containing medium. How would you generate an Hfr strain that is resistant to streptomycin and can metabolize lactose (Hint: F factors occasionally integrate into the chromosome to become Hfr strains, and occasionally Hfr strains excise their DNA from the chromosome to become F + strains that carry an F factor.)
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In your laboratory, you have an F strain of E. coli that is resistant to streptomycin and is unable to metabolize lactose, but it can metabolize glucose. Therefore, this strain can grow on a medium that contains glucose and streptomycin, but it cannot grow on a medium containing lactose. A researcher has sent you two E. coli strains in two separate tubes. One strain, let's call it strain A, has an F' factor (an F prime factor) that carries the genes that are required for lactose metabolism. On its chromosome, it also has the genes that are required for glucose metabolism. However, it is sensitive to streptomycin. This strain can grow on a medium containing lactose or glucose, but it cannot grow if streptomycin is added to the medium. The second strain, let's call it strain B, is an F strain. On its chromosome, it has the genes that are required for lactose and glucose metabolism. Strain B is also sensitive to streptomycin. Unfortunately, when strains A and B were sent to you, the labels had fallen off the tubes. Describe how you could determine which tubes contain strain A and strain B.
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Briefly describe the lytic and lysogenic cycles of bacteriophages. In your answer, explain what a prophage is.
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An Hfr strain that is hisE + and pheA + was mixed with a strain that is hisE and pheA . The conjugation was interrupted and the percentage of recombinants for each gene was determined by streaking on a medium that lacked either histidine or phenylalanine. The following results were obtained: img A. Determine the map distance (in minutes) between these two genes. B. In a previous experiment, it was found that hisE is 4 minutes away from the gene pabB. PheA was shown to be 17 minutes from this gene. Draw a genetic map describing the locations of all three genes.
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In the experiment of Figure 7.1, Joshua Lederberg and Edward Tatum could not discern whether met + bio + genetic material was transferred to the met bio thr + leu + thi + strain or if thr + leu + thi + genetic material was transferred to the met + bio + thr leu thi strain. Let's suppose that one strain is streptomycin-resistant (say, met + bio + thr leu thi ) and the other strain is sensitive to streptomycin. Describe an experiment that could determine whether the met + bio + genetic material was transferred to the met bio thr + leu + thi + strain or if the thr + leu + thi + genetic material was transferred to the met + bio + thr leu thi strain. FIGURE 7.1 Experiment of Lederberg and Tatum demonstrating genetic transfer during conjugation in E. coli. When plated on a growth medium lacking amino acids, biotin, and thiamine, the met bio thr + leu + thi + or met + bio + thr leu thi strains were unable to grow. However, if they were mixed together and then plated, some colonies were observed. These colonies were due to the transfer of genetic material between these two strains by conjugation. Note: In bacteria, it is common to give genes a three-letter name (shown in italics) that is related to the function of the gene. A plus superscript ( + ) indicates a functional gene, and a minus superscript ( ) indicates a mutation that has caused the gene or gene product to be inactive. In some cases, several genes have related functions. These may have the same three-letter name followed by different capital letters. For example, different genes involved with leucine biosynthesis may be called leuA, leuB, leuC, and so on. In the experiment described in Figure 7.1, the genes involved in leucine biosynthesis had not been distinguished, so the gene involved is simply referred to as leu + (for a functional gene) and leu (for a nonfunctional gene). img
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What is an interrupted mating experiment What type of experimental information can be obtained from this type of study Why is it necessary to interrupt mating
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Discuss the advantages of the genetic analysis of bacteria and bacteriophages. Make a list of the types of allelic differences among bacteria and phages that are suitable for genetic analyses.
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Conjugation is sometimes called "bacterial mating." Is it a form of sexual reproduction Explain.
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As mentioned in solved problem S2, origins of transfer can be located in many different locations, and their direction of transfer can be clockwise or counterclockwise. Let's suppose a researcher conjugated six different Hfr strains that were thr + leu + ton s str r azi s lac + gal + pro + met + to an F strain that was thr leu ton r str s azi r lac gal pro met and obtained the following results: img Draw a circular map of the E. coli chromosome and describe the locations and orientations of the origins of transfer in these six Hfr strains.
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If you mix together an equal number of F + and F cells, how would you expect the proportions to change over time In other words, do you expect an increase in the relative proportions of F + or of F cells Explain your answer.
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In the experiment of Figure 7.1, a met bio thr + leu + thi + cell could become met + bio + thr + leu + thi + by a (rare) double mutation that converts the met bio genes into met + bio +. Likewise, a met + bio + thr leu thi cell could become met + bio + thr + leu + thi + by three mutations that convert the thr leu thi genes into thr + leu + thi +. From the results of Figure 7.1, how do you know that the occurrence of 10 met + bio + thr + leu + thi + colonies is not due to these types of rare double or triple mutations FIGURE 7.1 Experiment of Lederberg and Tatum demonstrating genetic transfer during conjugation in E. coli. When plated on a growth medium lacking amino acids, biotin, and thiamine, the met bio thr + leu + thi + or met + bio + thr leu thi strains were unable to grow. However, if they were mixed together and then plated, some colonies were observed. These colonies were due to the transfer of genetic material between these two strains by conjugation. Note: In bacteria, it is common to give genes a three-letter name (shown in italics) that is related to the function of the gene. A plus superscript ( + ) indicates a functional gene, and a minus superscript ( ) indicates a mutation that has caused the gene or gene product to be inactive. In some cases, several genes have related functions. These may have the same three-letter name followed by different capital letters. For example, different genes involved with leucine biosynthesis may be called leuA, leuB, leuC, and so on. In the experiment described in Figure 7.1, the genes involved in leucine biosynthesis had not been distinguished, so the gene involved is simply referred to as leu + (for a functional gene) and leu (for a nonfunctional gene). img
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