Biochemistry
Biochemistry
6th Edition
ISBN: 9781305577206
Author: Reginald H. Garrett, Charles M. Grisham
Publisher: Cengage Learning
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Chapter 28, Problem 1P

Semiconservative or Conservative DNA Replication If 15 N -Iabeled E. coli DNA has a density of 1.724 g/mL, 14 N -labeled DNA has a density of 1.710 g/mL, and E. coli cells grown for many generations on 1 4 NH 4 + as a nitrogen source are transferred to media containing 1 5 NH 4 + as the sole N-source,

(a) What will be the density of the DNA after one generation, assuming replication is semiconservative?

(b) Suppose replication took place by a conservative mechanism in which the parental strands remained together and the two progeny strands were paired. Design an experiment that could distinguish between semiconservative and conservative modes of replication.

Expert Solution
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Interpretation Introduction

(a)

Interpretation:

The density of DNA of E.coli cells should be calculated after one generation that are grown for many generations on14NH4+ and then transferred to a media containing15NH4+ as only source of nitrogen assuming the replication is semiconservative given the density of15N-labeled E.coli DNA is 1.724 g/ml, density of14N-labeled E.coli DNA is 1.710g/ml.

Concept Introduction:

In model of semiconservative mode of replication parent DNA strands unwind and each one of these acts as a template for a new additional strand, thus synthesizes two DNA each with a pair of an old and a new strand.

Answer to Problem 1P

The average density after one generation of the transfer of media will be 1.717 g/ml.

Explanation of Solution

The density of DNA after one generation is calculated with arithmetic mean of densities of15N-labeled E.coli DNA (heavy strand) and the14N- labeled DNA (light strand).

Average density =ρ=(1.724+1.710)2=1.717g/ml

After the transfer of E.coli cells from a light medium of14NH4+ to a heavy medium of15NH4+, the cells have15NH4+ as only source of nitrogen. If a semi-conservative replication is assumed, replication produces two DNA each having an old strand of15N-labeled E.coli (ρ=1.724 g/ml ) and a new strand of14N- labeled E.coli.

Expert Solution
Check Mark
Interpretation Introduction

(b)

Interpretation:

Assume a replication by a conservative mechanism in which parental strands stayed together and two daughter strands paired. A brief description of an experiment that can differentiate between semiconservative and conservative modes of replication should be given.

Concept Introduction:

For the DNA strands to differentiate there needs to be a system of tagging the parent DNA. This is done by changing the medium of the E.coli cells. Cells that are grown in15NH4+ medium are transferred to a medium containing heavy nitrogen (14N)

Answer to Problem 1P

Grow E.coli cells for many generations in a medium containing15NH4+ as a source of nitrogen. Extract DNA of the cells and calculate its density using “CsCSl” density gradient centrifugation. Feed E.coli for many generations on14NH4+ medium and Similarly determine its density.

Now, transfer the cells grown on15NH4+ medium to the media containing14N. The number of cells is carefully monitored by colony formation assay. The DNA is then extracted generation after generation. Extract the DNA after first generation and determine its density.

Explanation of Solution

The pair of progeny DNA produced has an equal density intermediate between the first group and the second group. This is not the case with conservative replication as density of progeny DNA is not equal.

In model of conservative replication, the pair of progeny DNA has one DNA containing two helical old strands and the other is newly fabricated. In contrast the semiconservative model has same density for both progeny DNA, intermediate between the14N and15N E.coli. The result fits with the latter model.

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49 N), heavy parental DNA, and all nucleotides with 14N (another isotope of N) as "nucleotide supply source" for DNA replication in a tube. Use the figure provided below to illustrate the arrangement of light and heavy isotopes of nitrogen in DNA molecules formed in Generation one, two and three. (Please use the shade or different color to represent the daughter strands of DNA with light N-isotope, so as to be different from the parental DNA strands.) Make sure to label what each color represents clearly. Otherwise, the answer is not considered correct. semiconservative replication ப Generation Zero (Original DNA) Generation One Generation Two
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