Assume that a culture of E. coli was grown for approximately 50 generations in 15N (provided in the medium in the ammonium ion), which is a heavy isotope of nitrogen (14N). You extract the DNA from the culture, and it has a density of 1.723 gm/cm3 (water = 1.00 gm/cm3). From the literature, you determine that DNA containing only the common form of nitrogen, 14N, has a density of 1.700 gm/cm3. Bacteria from the 15N culture were washed in buffer and transferred to 14N medium for one generation immediately after which the DNA was extracted and its density determined.
(a) What would be the expected density of the extracted DNA?
(b) After you heat the extracted DNA until it completely denatures (95°C for 15 minutes), what would you expect the density of the DNA in the denatured extract to be? For the purposes of this question, assume that DNA has the same density regardless of whether it is single- or double-stranded.
(c) Assuming that the molar percentage of adenine in the extracted DNA was 20 percent, what would be the expected molar percentages of the other nitrogenous bases in this DNA?
(d) Assume that a fraction of the extracted DNA was digested to completion with the enzyme snake venom diesterase. This enzyme cleaves between the phosphate and the 3' carbon. Present a "simplified" diagram that would illustrate the structure of the predominant resulting molecule.
Culture and sensitivity
Escherichia coli (E. coli).
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Cell Grown in 15N and 14 N: Graph of Relative DNA Concentration Vs. the Density
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