{"id":244,"date":"2012-11-05T16:40:35","date_gmt":"2012-11-05T20:40:35","guid":{"rendered":"https:\/\/sites.owu.edu\/wolverton\/?page_id=244"},"modified":"2016-12-01T08:50:41","modified_gmt":"2016-12-01T13:50:41","slug":"dna-replication","status":"publish","type":"page","link":"https:\/\/sites.owu.edu\/wolverton\/teaching\/cell-bio\/dna-replication\/","title":{"rendered":"DNA Replication"},"content":{"rendered":"<p><!--:en--><\/p>\n<h2 id=\"links\">Links<\/h2>\n<p><a href=\"http:\/\/www.nature.com\/principles\/ebooks\/introduction-to-cell-biology-4570805\/4569936\">Principles Module 47<\/a><\/p>\n<h2 id=\"objectives\">Objectives<\/h2>\n<ul>\n<li>Explain how the structure of DNA suggests a mechanism for its replication<\/li>\n<li>Describe the design of the Meselson and Stahl experiment that demonstrated semiconservative replication<\/li>\n<li>Discuss the process of DNA replication<\/li>\n<li>Identify the key enzymes involved in replication and their functions<\/li>\n<\/ul>\n<h2 id=\"keypoints\">Key points<\/h2>\n<p>DNA structure<\/p>\n<ul>\n<li>each nucleotide is composed of a pentose sugar with a phosphate and a nitrogenous base<\/li>\n<li>early work showed A in equal amount as T, G equal to C<\/li>\n<li>X-ray diffraction studies by Franklin and Wilkins gave clues to structure<\/li>\n<li>Watson and Crick made the major conceptual breakthrough, proposing the double helix\n<ul>\n<li>used physical models made to scale of molecules to predict positions of atoms<\/li>\n<li>accounted for all observations on DNA to date, including equal quantities of bases<\/li>\n<li>suggested a mechanism of replication just based on structure<\/li>\n<li>watch an interview in which James Watson <a href=\"http:\/\/www.dnalc.org\/resources\/3d\/25-basepairing.html\" target=\"_blank\">describes the insight leading to the idea of complementary base pairing<\/a><\/li>\n<\/ul>\n<\/li>\n<\/ul>\n<div>Semiconservative Replication<\/div>\n<div>\n<ul>\n<li>Meselson and Stahl designed an experiment to test different models of replication using &#8216;heavy&#8217; N to label the nitrogenous base part of DNA\n<ul>\n<li>grew bacteria in 15N media for many generations, then transferred to 14N<\/li>\n<li>separated DNA by density gradient centrifugation<\/li>\n<li>different replication models made distinct, testable predictions:\n<ul>\n<li>Semiconservative replication predicts 1 density of DNA after 1 round, 2 after 2 or more rounds of replication<\/li>\n<li>Conservative replication predicts 2 densities of DNA after 1 or more rounds of replication<\/li>\n<li>Dispersive replication predicts 1 density after 1 or more rounds of replication<\/li>\n<\/ul>\n<\/li>\n<\/ul>\n<\/li>\n<\/ul>\n<div>Features of semiconservative replication<\/div>\n<div>\n<ul>\n<li>each strand of the double helix serves as a <a href=\"http:\/\/www.dnalc.org\/resources\/3d\/01-replication-the-helix.html\">template to produce a complementary strand<\/a>\u00a0(video link)<\/li>\n<li>each new DNA molecule is made up of one old, one new strand<\/li>\n<\/ul>\n<div>Overview of Replication<\/div>\n<div>\n<ul>\n<li>initiation: proteins open double helix, exposing single strands of DNA to enzymes<\/li>\n<li>elongation: DNA polymerases add free nucleotide bases in 5&#8242; to 3&#8242; direction<\/li>\n<li>termination: polymerases fall off template strands<\/li>\n<\/ul>\n<div>Replication in Prokaryotes in detail<\/div>\n<div>\n<ul>\n<li>begins at a specific sequence of DNA known as the origin of replication (oriC), rich in AT sequences<\/li>\n<li>DNA polymerase enzymes only add free nucleotides to the -OH on the 3&#8242; carbon, thus replication proceeds from 5&#8242; end to 3&#8242; end of polynucleotide\n<ul>\n<li>DNA polymerase III is the key enzyme for replication in prokaryotes<\/li>\n<\/ul>\n<\/li>\n<li>primase adds a short sequence of RNA &#8216;primer&#8217; to DNA template\n<ul>\n<li>primase does not require the free 3&#8242;-OH group to catalyze addition<\/li>\n<\/ul>\n<\/li>\n<li>DNA helicases unwind the double helix, require ATP<\/li>\n<li>once unwound, DNA binds to single-strand binding proteins (SSBs) that prevent re-pairing<\/li>\n<li>topoisomerases bind to double-stranded DNA and relieve torsional strain<\/li>\n<\/ul>\n<div>Leading and lagging strands<\/div>\n<div>\n<ul>\n<li>leading strand synthesis is toward the replication fork (where DNA is unwinding), can be replicated from a single primer<\/li>\n<li>lagging strand is away from replication fork, requires repeated priming events, which creates Okazaki fragments<\/li>\n<li>DNA replication on the lagging strand is discontinuous<\/li>\n<li>DNA polymerase I replaces RNA primers with DNA<\/li>\n<li>DNA ligase joins Okazaki fragments by forming phosphodiester bond<\/li>\n<\/ul>\n<div>Eukaryotic differences<\/div>\n<div>\n<ul>\n<li>larger genome of Eukaryotes requires multiple origins, forming distinct replicons<\/li>\n<li>the overall process is similar but involves more components<\/li>\n<li>Eukaryotic chromosomes are linear, not circular, presenting a problem for replication on the lagging strand at the end<\/li>\n<li>telomerase solves this problem by having a built-in RNA template for repeating telomere sequence<\/li>\n<li>keeps telomeres from shortening after each replication cycle<\/li>\n<\/ul>\n<\/div>\n<\/div>\n<\/div>\n<\/div>\n<\/div>\n<\/div>\n<h2 id=\"in-classactivities\">In-class activities<\/h2>\n<h2 id=\"questionsforpractice\">Questions for Practice<\/h2>\n<ul>\n<li>In what way did the structure of DNA proposed by Watson and Crick contain a mechanism for replication &#8216;built-in&#8217;?<\/li>\n<li>Describe the experiment that showed DNA replication was semi-conservative.<\/li>\n<li>What is the role of _________ (helicase, topoisomerase, primase, DNA polymerase III, polymerase I) in DNA replication?<\/li>\n<li>What is an Okazaki fragment?<\/li>\n<\/ul>\n<p><!--:--><\/p>\n","protected":false},"excerpt":{"rendered":"<p>Links Principles Module 47 Objectives Explain how the structure of DNA suggests a mechanism for its replication Describe the design of the Meselson and Stahl experiment that demonstrated semiconservative replication Discuss the process of DNA replication Identify the key enzymes involved in replication and their functions Key points DNA structure each nucleotide is composed of [&hellip;]<\/p>\n","protected":false},"author":40,"featured_media":0,"parent":447,"menu_order":0,"comment_status":"closed","ping_status":"closed","template":"","meta":{"footnotes":""},"class_list":["post-244","page","type-page","status-publish","hentry"],"_links":{"self":[{"href":"https:\/\/sites.owu.edu\/wolverton\/wp-json\/wp\/v2\/pages\/244","targetHints":{"allow":["GET"]}}],"collection":[{"href":"https:\/\/sites.owu.edu\/wolverton\/wp-json\/wp\/v2\/pages"}],"about":[{"href":"https:\/\/sites.owu.edu\/wolverton\/wp-json\/wp\/v2\/types\/page"}],"author":[{"embeddable":true,"href":"https:\/\/sites.owu.edu\/wolverton\/wp-json\/wp\/v2\/users\/40"}],"replies":[{"embeddable":true,"href":"https:\/\/sites.owu.edu\/wolverton\/wp-json\/wp\/v2\/comments?post=244"}],"version-history":[{"count":1,"href":"https:\/\/sites.owu.edu\/wolverton\/wp-json\/wp\/v2\/pages\/244\/revisions"}],"predecessor-version":[{"id":499,"href":"https:\/\/sites.owu.edu\/wolverton\/wp-json\/wp\/v2\/pages\/244\/revisions\/499"}],"up":[{"embeddable":true,"href":"https:\/\/sites.owu.edu\/wolverton\/wp-json\/wp\/v2\/pages\/447"}],"wp:attachment":[{"href":"https:\/\/sites.owu.edu\/wolverton\/wp-json\/wp\/v2\/media?parent=244"}],"curies":[{"name":"wp","href":"https:\/\/api.w.org\/{rel}","templated":true}]}}