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Chapter 16: The Molecular Basis of Inheritance Key Concepts 16.1 DNA is the genetic material 16.2 Many proteins work together in DNA replication and repair Interactive Question 6.1: Hershey and Chase devised an experiment using radioactive isotopes to determine whether the phage’s DNA or protein entered the bacteria and was the genetic material of T2 page. a. How did they label phage protein? Grew the T2 bacteriophage with E. coli that had radioactive sulfur b. How did they label phage DNA? Used radioactive phosphorous Separate samples of E. coli were infected with the differently labeled T cells, then blended and centrifuge to isolate the bacterial cells from he lighter viral particles. c. Where was the radioactivity found in the samples with labeled phage protein? Radioactivity was found in supematent—protein did not enter the phage d. In the samples with labeled phage DNA? Most of the radioactivity was in pellet boci cell e. What did Hershey and Chase conclude from these results? Concluded that viral DNA was injected into the bacterial cell and some as the hereditary material for viruses Interactive Question 16.2: Review the structure of DNA by labeling the following diagrams. a. sugar-phosphate backbone b. 3’ end c. pyrimidin d. purine e. hydrogen bond f. cytosine g. guanine h. adenine i. thymine j. 5’ end k. nucleotide l. deoxyribose m. phosphate group n. 3.4 nm o. 0.34 nm p. 02. nm Interactive Question 16.3: Using different colors for heavy (parental) and light (new) strands of DNA, sketch the results of two replication cycles when E. coli were moved from medium containing 15N to 14N medium. Show the resulting density bands in he centrifuge tubes. Interactive Question 16.4: Lock back to Interactive Question 16.2 and label the 5’ and 3’ ends of both strands of the DNA molecule. 5’ end = phosphate (PO4-3) hydroxyl group 3’ end = OH- (hydroxyl group) and sugar Interactive Question 16.5: In this diagram showing the replication of DNA, label the following terms: leading and lagging strands, Okazaki fragment, DNA pol III, DNA pol I, DNA ligase, helicase, primase, single-strand binding proteins, RNA primer, replication fork, and 5’ and 3’ ends of parental DNA. a. helicase (unwinds) b. SSBP’s (keeps strands separated) c. DNA polymerase III d. Leading strand e. Lagging strand f. DNA ligase (joins DNA) g. DNA polymerase I (replaces primers) h. Okazaki fragment i. j. k. l. m. RNA primer replication fork primase 3’ end 5’ end Structure Your Knowledge 1. Summarize the evidence and techniques Watson and Crick used to deduce the doublehelix structure of DNA. Watson and Crick used the X-ray diffraction photo of Franklin to deduce that DNA was a helix 2 nm wide, with nitrogenous bases stacked 0.34 nm apart, and making a full turn every 3.4 nm. Franklin had concluded that the sugar-phosphate backbones were on the outside for the helix with the bases extending inside. Using molecular models of wire, Watson and Crick experimented with various arrangements and finally paired of a purine base with a pyrimidine base, which produced the proper diameter. Specificity of base pairing (A with T and C with G) is assured by hydrogen bonds. 2. Review your understanding of DNA replication by describing key enzymes and proteins in the order of their functioning) that direct replication. Replication bubbles form where proteins recognize specific base sequences and open up the two strands. Helicase, an enzyme that works at the replication fork, untwists the helix and separates the strands. Topoisomerase eases the twisting ahead of the replication for. Single-strand binding proteins support the separated strands while replication takes place. Primase lays down about 10 RNA bases to start the new strand. After a proper base pairs up on the exposed template, DNA polymerase III joins the nucleotide to the 3’ end of the new strand. On the lagging strand, short Okazaki fragments are formed by primase and polymerase (again moving 5’3’). DNA polymerase I replaces the primer. Ligase joins the 3’ end of one fragment to the 5’ end of its neighbor. Proofreading enzymes check for mispaired bases, and nucleases, DNA polymerase, and other enzymes repair damage or mismatches. Test Your Knowledge 1. One of the reasons most scientists believed proteins were the carriers of genetic information was that c. proteins were much more complex and heterogeneous molecules than nucleic acids. (pg. 293) 2. Transformation involves a. the uptake of external genetic material, often from one bacterial strain to another. (pg. 294) 3. The DNA of an organism has thymine as 20% of is bases. What percentage of its bases would be guanine? b. 30% (pg. 298) 4. In his work with pneumonia-causing bacteria, Griffith found that: d. some heat-stable chemical was transferred to harmless cells to transform them into pathogenic cells. (pg 294) 5. When T2 pages are grown with radioactive sulfur, b. their proteins are tagged. (pg 294-295) 6. Meselson and Stahl a. provided evidence for the semiconservative model of DNA replication. (pg. 299-300) 7. Watson and Crick concluded that each base could not pair with itself because b. the uniform width of 2 nm would not permit two purines or two pyrimidines to pair together. (pg. 298) 8. The joining of nucleotides in the polymerization of DNA requires energy from e. they hydrolysis of the pyrophosphates removed from nucleoside triphosphates. (pg 300-301) 9. Continuous elongation of a new DNA strand along one strand of DNA c. occurs on the leading strand. (pg. 301) 10. Which of the following statements about DNA polymerase is incorrect? a. It forms the bonds between complementary base pairs. (pg. 301) 11. Thymine dimers—covalent links between adjacent thymine bases in DNA—may be induced by UV light. When they occur, they are repaired by e. a, b, and c are all needed. (pg. 306) 12. How does DNA synthesis along the lagging strand differ from that on the leading strand? d. Okazaki fragments, which each grow 5’ 3’, must be joined along the lagging strand. (pg. 302) 13. Which of the following enzymes or proteins is paired with an incorrect or inaccurate function? e. Primase—form DNA primer to start replication (pg. 303) 14. Which letter indicates the 5’ end of this single DNA strand? a. (pg. 296) 15. At which letter would the next nucleotide be added? d. (pg. 296) 16. Which letter indicates a phosphodiester bond formed by DNA polymerase? b. (pg. 296) 17. The base sequence of the DNA strand made from this template would be (from top to bottom c. T A C. (pg. 296) 18. T2 phage is grown in E. coli with radioactive phosphorus and then allowed to infect other E. coli. The culture is blended to separate the viral coats from the bacterial cells and centrifuged. Which of the following best describes the expected results of such an experiment? d. Viral DNA is labeled; radioactivity is found in the pellet. (pg 294-295) 19. What are telomeres, and what do they do? e. Both a and b are correct. (pg. 306) 20. You are trying to support your hypothesis that DNA replication is conservative; ie., parental strands are separate complementary strands are made, but these new strands join together to make a new DNA molecule, and the parental strands rejoin. You take E. coli that had grown in a medium containing only heavy nitrogen (15N) and transfer a sample to a medium containing light nitrogen (14N). After allowing time for only one DNA replication, you centrifuge a sample and compare the density band(s) formed with control bands for bacteria grown on ether normal 14N or 15N medium. Which band location would support your hypothesis of conservative DNA replication? c. (pg. 300) 21. Using the experiment explained in question 20, which centrifuge tube would represent the band distribution obtained after one replication showing that DNA replication is semiconservative? a. (pg. 300)