Molecular Biology of the Gene Chapter 10 Molecular Biology of the Gene

DNA REPLICATION © 2012 Pearson Education, Inc. 2

10.4 DNA replication depends on specific base pairing In their description of the structure of DNA, Watson and Crick noted that the structure of DNA suggests a possible copying mechanism. DNA replication follows a semiconservative model. The two DNA strands separate. Each strand is used as a pattern to produce a complementary strand, using specific base pairing. Each new DNA helix has one old strand with one new strand. Student Misconceptions and Concerns The authors note that although the general process of semiconservative DNA replication is relatively simple, it involves complex biochemical gymnastics. The DNA molecule is unwound, each strand is copied simultaneously, the correct bases are inserted, and the product is proofread and corrected. Before discussing these details, be sure that your students understand the overall process, what is accomplished, and why each step is important. Teaching Tips 1. Demonstrate the complementary base pairing within DNA. Present students with the base sequence to one side of a DNA molecule and have them work quickly at their seats to determine the sequence of the complimentary strand. For some students, these sorts of quick practice are necessary to reinforce a concept and break up a lecture. 2. The semiconservative model of DNA replication is like making a photo from a negative and then a new negative from the photo. In each new negative and photo pair, the new item was made from an old item. Animation: DNA Replication Overview © 2012 Pearson Education, Inc. 3

A parental molecule of DNA Figure 10.4A_s1 A T C G G C A T T A A parental molecule of DNA Figure 10.4A_s1 A template model for DNA replication (step 1) 4

A parental molecule of DNA Figure 10.4A_s2 A T T A A T C G C G G C G C G C C A A T A T Free nucleotides T A T A A parental molecule of DNA The parental strands separate and serve as templates Figure 10.4A_s2 A template model for DNA replication (step 2) 5

A parental molecule of DNA Figure 10.4A_s3 A T T A A T A T A T C G C G G C G C G C G C G C C G C G C A A T A T A T A T Free nucleotides T A T A T A T A A parental molecule of DNA The parental strands separate and serve as templates Two identical daughter molecules of DNA are formed Figure 10.4A-s3 A template model for DNA replication (step 3) 6

Problems and Solutions Associated with DNA Replication Replication must occur quickly. Multiple origin of replication sites are started along eukaryotic chromosomes. DNA polymerase catalyzes DNA synthesis Parental DNA molecule Origin of replication “Bubble” Parental strand Daughter strand Two daughter DNA molecules Student Misconceptions and Concerns The authors note that although the general process of semiconservative DNA replication is relatively simple, it involves complex biochemical gymnastics. The DNA molecule is unwound, each strand is copied simultaneously, the correct bases are inserted, and the product is proofread and corrected. Before discussing these details, be sure that your students understand the overall process, what is accomplished, and why each step is important. Teaching Tips 1. To explain the adaptive advantage of multiple replication sites over a single site of replication, ask the students to imagine copying, by hand, the first ten chapters of your biology textbook. The task would certainly go faster if ten students each copied a different chapter. 2. There are about 500,000 words in the Biology: Concepts & Connections textbook. The accuracy of DNA replication would be like copying every word in this textbook by hand 2,000 times and writing just one word incorrectly, making one error in every 1 billion words. Animation: Origins of Replication © 2012 Pearson Education, Inc. 7

Problems and Solutions Associated with DNA Replication DNA strands are twisted around each other and must be separated Helicase Enzyme that unwraps DNA and separate H-bonds between strands Student Misconceptions and Concerns The authors note that although the general process of semiconservative DNA replication is relatively simple, it involves complex biochemical gymnastics. The DNA molecule is unwound, each strand is copied simultaneously, the correct bases are inserted, and the product is proofread and corrected. Before discussing these details, be sure that your students understand the overall process, what is accomplished, and why each step is important. Teaching Tips 1. To explain the adaptive advantage of multiple replication sites over a single site of replication, ask the students to imagine copying, by hand, the first ten chapters of your biology textbook. The task would certainly go faster if ten students each copied a different chapter. 2. There are about 500,000 words in the Biology: Concepts & Connections textbook. The accuracy of DNA replication would be like copying every word in this textbook by hand 2,000 times and writing just one word incorrectly, making one error in every 1 billion words. © 2012 Pearson Education, Inc. 8

Problems and Solutions Associated with DNA Replication Replication must be accurate DNA polymerase can proof-read its own work Correct mistakes Student Misconceptions and Concerns The authors note that although the general process of semiconservative DNA replication is relatively simple, it involves complex biochemical gymnastics. The DNA molecule is unwound, each strand is copied simultaneously, the correct bases are inserted, and the product is proofread and corrected. Before discussing these details, be sure that your students understand the overall process, what is accomplished, and why each step is important. Teaching Tips 1. To explain the adaptive advantage of multiple replication sites over a single site of replication, ask the students to imagine copying, by hand, the first ten chapters of your biology textbook. The task would certainly go faster if ten students each copied a different chapter. 2. There are about 500,000 words in the Biology: Concepts & Connections textbook. The accuracy of DNA replication would be like copying every word in this textbook by hand 2,000 times and writing just one word incorrectly, making one error in every 1 billion words. © 2012 Pearson Education, Inc. 9

Problems and Solutions Associated with DNA Replication DNA Polymerase can only add nucleotides to the 3’ end of a pre-exiting DNA strand One-strand synthesized as a single continuous strand (leading strand) The other (lagging strand) synthesized in small fragments (Okazaki fragments) DNA Ligase connects these fragments RNA polymerase (primase) starts replication by building a small, RNA primer Student Misconceptions and Concerns The authors note that although the general process of semiconservative DNA replication is relatively simple, it involves complex biochemical gymnastics. The DNA molecule is unwound, each strand is copied simultaneously, the correct bases are inserted, and the product is proofread and corrected. Before discussing these details, be sure that your students understand the overall process, what is accomplished, and why each step is important. Teaching Tips 1. To explain the adaptive advantage of multiple replication sites over a single site of replication, ask the students to imagine copying, by hand, the first ten chapters of your biology textbook. The task would certainly go faster if ten students each copied a different chapter. 2. There are about 500,000 words in the Biology: Concepts & Connections textbook. The accuracy of DNA replication would be like copying every word in this textbook by hand 2,000 times and writing just one word incorrectly, making one error in every 1 billion words. © 2012 Pearson Education, Inc. 10

DNA strands run in opposite directions. 5 end 3 end 5 4 3 2 1 P HO A T C G OH Remember….. DNA strands run in opposite directions. 5’ indicates open P end 3’ indicates open -OH end Figure 10.5B The opposite orientations of DNA strands 11

DNA Polymerase can only attach nucleotides to 3’-end New strand Template strand 5¢ end 3¢ end 5¢ end 3¢ end Sugar Base Phosphate DNA polymerase 3¢ end 3¢ end Pyrophosphate Nucleoside triphosphate 5¢ end 5¢ end

Animation: Leading Strand Animation: Lagging Strand 3 DNA polymerase molecule This daughter strand is synthesized continuously 5 Parental DNA 5 3 Replication fork This daughter strand is synthesized in pieces 3 5 Animation: Leading Strand 5 3 Figure 10.5C How daughter DNA strands are synthesized Animation: Lagging Strand DNA ligase Overall direction of replication 13

Overall direction of replication LE 16-14 3¢ Parental DNA 5¢ Leading strand 5¢ 3¢ Okazaki fragments Lagging strand 3¢ 5¢ DNA pol III Template strand Leading strand Lagging strand Template strand DNA ligase Overall direction of replication

Animation: DNA Replication Review Overall direction of replication Leading strand Lagging strand Origin of replication Lagging strand Leading strand OVERVIEW DNA polymerase Leading strand DNA ligase Replication fork 5¢ DNA polymerase 3¢ Primase DNA polymerase Parental DNA Lagging strand Primer 3¢ 5¢ Animation: DNA Replication Review

Bacteria cultured in medium containing 15N Bacteria transferred to medium containing 14N DNA sample centrifuged after 20 min (after first replication) DNA sample centrifuged after 40 min (after second replication) Less dense More dense First replication Second replication Conservative model Semiconservative model