You digest a 5 kb plasmid with two enzymes that yield fragments of 4 kb and 1 kb. After running the digest on a gel, what would you predict to see in terms of the intensities of the bands?
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- You want to digest 1 µg of plasmid DNA in a final volume of 50 μL. Your solution containing plasmid has a concentration of 25 ng/μL. How many μL of your plasmid solution do you need to add to your reaction tube to digest your desired mass of plasmid?You're purifying some plasmid DNA from a culture of bacteria and you want to know how pure it is. You measure the optical density at 260 nm and 280 nm and find the ratio is 2.0. You suspect there is RNA contamination in your preparation, so you treat your preparation with RNase. But the ratio is still 2.0. Protein assays tell you there is no protein in your solution, and no other biological molecules absorb light very efficiently at those wavelengths. What's the explanation?You are about to isolate a 3000 bp large plasmid from an E.coli culture. You know that the plasmid is present in 100 copies per E. coli cell. You aim to have a final plasmid concentration of 100 ng/µl in a total volume of 50 ul. Assuming the yield is 100 %, how many E. coli cells should the culture from which the plasmid is to be isolated at least contain A) 4.7 · 1013 cells B) 1.6 · 1010 cells C) 3.2 · 1010 cells D) 3.2 · 1026 cells E) 1.6 · 1012 cells Solution B that was used during the plasmid isolation contains 0.2 M NaOH (see practical manual). Which effect does NaOH have on E. coli DNA A) It denatures genomic DNA and plasmid DNA. B) It denatures genomic DNA and but not plasmid DNA. C) It denatures plasmid DNA but not genomic DNA. D) It denatures neither genomic DNA nor plasmid DNA. E) This is unpredictable. Which statement on the migration of DNA fragments through agarose gels is false A) Small fragments migrate faster than larger fragments because they can move faster through…
- You have begun your career in medicinal biochemistry and have just discovered a bacterial DNA plasmld (transferabl ring of DNA) that appears to destroy the Ebola virus. In order to characterize your new plasmid, the molar mass of the plasmid must be determined. You dissolve 25.00 mg of the purified plasmid in 0.200 mL of water at 2 °C and find the osmotlc pressure of this solution is 1.20 Torr at 20 °C and 1 atm pressure. Answer the following about the Ebola-killing plasmid. 33.) The osmotlc pressure of the system is: (a) 1 atm (b) 0.016 atm (c) 6.5 X 10-5 atm (d) 22.59 atm (e) 0.0016 atmYou're purifying some plasmid DNA from a culture of bacteria and you want to know how pure it is. You measure the optical density at 260 m and 280 m and find the ratio is 2.0. You suspect there is RNA contamination in your preparation, so you treat your preparation with RNase. But the ratio is still 2.0. Protein assays tell you there is no protein in your solution, and no other biological molecules absorb light very efficiently at those wavelengths. What's the explanation?You are studying a new plasmid, and you digest the plasmid with three restriction enzymes: Eco RI (E), Hindlll (H), and Xbal (X). You digest the plasmid DNA with each of the following combinations of enzymes and observe the results on an agarose gel. You are provided a partial plasmid map as shown below to the right. +18 a. What is the size of this plasmid in base pairs? b. What is the distance in base pairs between E1 and H? c. What is the distance in base pairs between E1 and X? d. What is the distance in base pairs between E2 and H? e. What is the distance in base pairs between E2 and X?
- Consider the following plasmid (size 8000 bp), with restriction sites at the positions indicated: (see image) a) This plasmid is digested with the enzymes listed below. Indicate how many fragments will begenerated in each case, and give the sizes of the fragments.PstIXhoICombination of PstI + XhoI + EcoRI (triple digest) b) Draw the banding pattern you would expect to observe if each of these digestions is loaded into a separate well of an agarose gel, and the fragments separated by electrophoresis. In the first well you load a DNA marker (M) containing fragments with sizes of 1000 bp, 2000 bp, 4000 bp and 8000 bp. c) This gel is transferred to a membrane in a Southern blot experiment, and hybridised to a radioactively labelled 200 bp probe, which anneals to the plasmid DNA at the position indicated on the diagram above. Draw the autoradiographic profile you would expect to observe for the membrane.a) A plasmid DNA in bacteria has a length of 14,000 bp and an Lk of 1300. Calculate the superhelical density o for this plasmid. Show your work for partial credit, round to one digit after the decimal point. b) You use a Type II topoisomerase to change the linking number of this plasmid to 1310. How many turnovers must the topoisomerase perform? Is this resulting plasmid underwound or overwound?The competency of bacterial cells to take up plasmids from the environment can be enhanced by treating them with calcium chloride. Which of the following statements is true regarding this process? Question 23 options: Chloride ions neutralize charges on the phospholipids and DNA. Chloride ions adhere to the cell membrane and calcium ions to the plasmid DNA, thus increasing the attractive force between them. The calcium ions change the structure of the cell membrane and, as a result, the pores are enlarged. Calcium ions neutralize charges on the phospholipids of the bacterial cell membrane and on the DNA of the plasmid. Chloride ions enter the cell through protein pores in the membrane, carrying plasmid DNA with them.
- The transformation results below were obtained with 10 ul of intact plasmid DNA at nine concentrations. The following numbers of colonies are obtained when 100 ul of transformed cells are plated on selective medium: Fill in the following table: Concentration # colonies DNA mass of Fraction of Mass Transformation PGREEN (Concentration x volume OR X spread = x 10ul plasmid solution) PGREEN in cell Cell efficiency Y÷ A suspension suspension spread = 100 ul - total vol cell susp. (Colonies - Mass spread) C x Z = A See (510 ul) HINT: this calculation is constant Given= X Given=Y С. Z. 0.00001 ug/ul | 4 0.00005 ug/ul 12 0.0001 ug/ul 0.0005 ug/ul 32 125 0.001 ug/ul 442 0.005 µg/ul 0.01 ug/ul 0.05 ug/ul 0.1 ug/ul 542 507 475 516 0.5 ug/ul 505The BamH1 enzyme comes at a concentration of 100,000 U/ml. You are asked to digest 20 ug of DNA with this enzyme. Determine: a) How many units will you need? b) How will you dispense them?You have two cell cultures, each containing a different plasmid. The first plasmid is ~5kbp long and contains three 5'-TCGA-3' and the other is the same size, but only contains two 5'-TCGA-3' sequences. You forget to label the two cell cultures you grew and have to figure out which one contains each plasmid. How would you go about identifying the the two cultures.
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