BIOL-L 211 Lecture 8 Outline of Last Lecture I. ArticlesII. HistonesIII. The NucleosomeOutline of Current Lecture I. Replication HypothesesII. Meselson-Stahl ExperimentIII. Methods and Minutia of DNA ReplicationCurrent LectureReplication II. Replication HypothesesA. Dispersive Model: Proposed that parental DNA strands fragmented and then served as templatesB. Semiconservative Model: Proposed that DNA "unzipped," yielding two identical template strands which then served as templates for new DNA1. Watson and Crick (from earlier lectures) preferred this modelC. Conservative Model: parental strands of DNA remained together throughout replicationII. Meselson-Stahl ExperimentA. Purpose: Discover mechanism for DNA replicationB. Procedure:1. Bacteria grown in media with nucleotide precursors labeled with a heavy isotope of Nitrogen (15N)2. Bacteria then transferred to media with 14N precursors (light isotope)These notes represent a detailed interpretation of the professor’s lecture. GradeBuddy is best used as a supplement to your own notes, not as a substitute.3. Bacteria allowed to grow in 14N solution4. Three samples placed in ultracentrifuge through a CsCl gradienta. DNA now separated by density5. ObserveC. Results: After two generations, the semiconservative model proved to be accurateIII. Methods and Minutia of DNA ReplicationA. Origins of Replication1. Prokaryotes have ONE2. Eukaryotes have hundreds to thousands3. Pre-Replication Complex: melts DNA strands apart at origins of replicationa. Specialized proteins find the origins of replicationB. The Replication Fork1. DNA synthesizes in two directions: toward and away from the replication fork (but still 5'->3' both ways)a. Leading Strand: synthesized toward/with the replication forkb. Lagging Strand: synthesized piece (Okazaki fragment) by piece in the direction opposite the replication fork2. Helicases: rings that unwind DNA (requiring ATP hydrolysis)3. Primases: enzymes that add primers to single stranded DNAa. Primers: starting point for DNA synthesis4. Primer:template junction: Combination of primer and template stranda. Template Strand: DNA is synthesized based off of this strand5. DNA polymerase: enzyme that synthesizes DNA (it's like a hand!!)6. Topoisomerase: enzyme that relieves structural tensiona. Resolves positive supercoils: knot of DNA that builds up in front of replication forkb. Creates instead negative supercoils: which are a natural and good conformation of DNA7. Single-stranded DNA binding proteins (SSBs): bind newly separated DNA strands and keep ssDNAs apartC. DNA Polymerase (hand)1. Palm: catalyzes DNA synthesis and monitors base pairing2. Fingers: bend template strand and set up next base to be paired3. Thumb: ensures that DNA polymerase does not dissociate from DNAD. Deoxynucleoside Triphosphate (dNTP): dATP, dGTP, dCTP, dTTP1. Base pair connected to triphosphate group via a deoxyribose group2. The phosphates are labeled alpha, beta, and gamma respectively with regards to their proximity to the nucleoside3. To form a phosphodiester bond, the beta and gamma phosphates are removed (post attack of the alpha phosphate by the 3' OH of the primer)4. The template strand controls which dNTPs are
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