Class Note for ECOL 320 at UA
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Date Created: 02/06/15
DNA Replication A CG39 CG gt Parent duplex GC Template strands Replica strands Reminder 39 Genes are DNA 0 DNA is a double stranded molecule 0 The genetic information in a gene is encoded 1n the sequence of bases on one strand of 1quot 1 10 20 3O 4O 50 6O 7O 80 90 100 AcatttgcttctgacacaactgtgttcactagcaactcaaacagacaccATGGTGCACCTGACTCCTGAGGAGAAGTCTGCCGTTACTGCCCTGTGGGGC 101 AAGGTGAACGTGGATGAAGTTGGTGGTGAGGCCCTGGGCAthtggtatcaaggttacaagacaggtttaaggagaccaatagaaactgggcatgtggag 201 acagagaagactcttgggtttctgataggcactgactctctctgcctattggtctattttcccacccttagGCTGCTGGTGGTCTACCCTTGGACCCAGA When a cell divides both daughter cells must receive a complete set of genes so the DNA molecules chromosomes must replicate before diVision Asexual Reproduction Review from Introductory Biology Prokaryotes l r i i rim l i i I 39i J repiication partitioning l The entire genome is on one circular chromosome 2 DNA molecule 2 The chromosome replicates once to produce two chromosomes that are identical except for rare mutations 3 The two identical daughter chromosomes move toward opposite end of the cell 4 When the cell divides the daughter chromosomes are partitioned one to each daughter cell Asexual Reproduction cont Eukaryotes Asexual reproduction by mitosis Cell Cycle Mitosis 62 Q G1 G1 S 62 interphase S Variable lengths Total time 15 m inutes gt days Animal cells in culture ca 1 day DNA replicates during S Gene expression occurs during G1 and G2 and S Nuclear division mitosis occurs during Mitosis Cell division cytokinesis occurs at the end of Mitosis Mitosis continued f I x x M NH frag 39 J quot W x x r TO r 39 rquot 9a H T I la X A rid 233 5 x h i i V I H 0 j I a 3 392 IL I F xxx 2 39x Q 2 53 P I X e quot39 ff K O oP k HL JFFF 39 39 Ld quota quot LJ J 39 The genome is divided among a number of chromosomes 1 Each chromosome replicates once in the S phase to produce two sister chromatids identical DNA molecules 2 During mitosis the the kinetochore regions of each pair of sister chromatids are attached by chromosome fibers to opposite poles of the cell 3 Chromosome fibers contract pulling sister chromatids to opposite ends of the cell 4 During cytokinesis the sister chromatids are partitioned one to each daughter cell Asexual Reproduction a Q 31 i Q i l i 17 X ifquot j replication partitioning r X39F K l 4 ah P Note that the end result of asexual reproduction in prokaryotes and eukaryotes is the same genetically 0 Each daughter cell gets a complete copy of the parental cell genome 0 The daughter cells are genetically identical except for neW mutations that occur during the cell cycle mainly during DNA replication 0 The daughter cells constitute a clone H l a l E I V b l I J I i DNA Replication is Semiconservative The strands separate A new strand is made using each 01d strand as a template according to the rules of base pairing Model proposed by Watson and Crick verified by Matt Meselson and Frank Stahl A r TA CG39 QT Parent duplex Replica strands DNA Replication Enzyme Activities Many enzymes are required for DNA replication We will only consider enzyme activities not specific enzymes Enzymes with these activities are also used for DNA manipulation in the lab Arthur Kornberg 1 Helicase unwinds double helical DNA 2 Singlestrand binding protein binds single strand to keep DNA unwound 3 DNA polymerase adds new nucleotides nucleoside triphosphates to 3 end of existing DNA strand or RNA primer Elongates chains 5 to 3 only 539 339 3 DNA Replication Enzyme Activities 4 Primase makes a single strand of ca 20 bp off RNA using a DNA strand as a template gt 3 539 t 3 W Reminder RNA is like DNA except Osingle stranded Oribose instead of deoxyribose ouracil instead of thymine U pairs with A just as T does 9 HE H CH OH CH OH H c o H c o 2 o 2 o C C c 0 H H H H N N H H H H H c H H c H OH OH H H o o Deoxyribose Ribose Thymine Uracil DNA Replication Enzyme Activities 5 Exonuclease removes nucleotides from the end of a DNA strand different enzymes work 5 to 3 or 3 to 5 1 11111111111 6 Ligase joins ends of single DNA strands by making neW phosphate bonds 1111mm Putting it All Together 1 primase makgs Rpm primer qum I M 539 rmwmm pmgmerase extenda 1 1 Putting it All Together 2 exunude ase remo was Firm pn mer 39539 rmnnmnmrrrrrmrrmmmnmrmmmmrrrrmmr pnlgmerasa fins gap l quot39 539 Emmi ligasa seals nick H I 539 mm Replicating Linear Chromosomes origin 1 l 1 539 mm 3 5 1 1 2 1 1 339 km 539 2 1 1 2 i F When Replication Forks Meet Replication Replication begins and is origin gt bidirectional Vi V WWW X eqummwmomm W N39 Wewa Replication origin l mmmmwxmqml mecml will MIL y lmmmmmmww Thwmm Synthesis starts at second origin Replication bubbles nal is also fuse where they meet bldlrectlonal xxx Newme WWW qu m0 1 1 comm lt gt W L 1404 lL ul J J ibxvxmx xmxmmmxmmm mw Enzyme Activities to Finish the Job 7 Gyrase a topoisomerase relaxes supercoils produced When the molecule is twisted during replication Also facilitates unwinding at beginning of replication 8 Telomerase uses a short RNA template to add short DNA repeats to the short ends of linear chromosomes When the last primer is removed using RNA template 5 5 5 l primer 4 l 3 v MSI CCCTI39CCC39ITCCC39I39I39 Enzyme Activities for Biotechnology These enzyme activities plus a few others are also used to manipulate DNA for example 0 PCR 0 Making recombinant DNA 0 Detecting mutations at the molecular level DNA Replication Details Polymerase Direction of strand growth OH G T Incoming nucleotl e FL lt51 c393 H Released Growing 339 end of new strand Template strand DNA Replication Details Lagging and Leading Strands Gyrase Cleaves and RNA primer initiates new strand synthesis DNA ligase joins Okazaki fragments on lagging strand swivels duplex to release mechanical stress of unwinding Helicase Gyrase topoisomerase II Helicase unwinds DNA at the replication fork Singlestrand binding proteins 888 proteins DNA polymerase DNA ligase complex Primosome DNA polymerase complex SSB proteins bind and stabilize single stranded DNA at replication fork Replicating Circular DNA Molecules 0 Bidirectional Replication Replication Replication 39 39 origin Elegll39llgatlon ongm i 7 7 Replication direction if both directions onl r l 39 mNmultaneously y M H1 3 AW 3 11 C Y U39d39 H B39d39 r I C 7 5 54223523 iwM n Iquot g 2 L A Replication g i 1 Replication fork Replicationlhmm fork 1123 WINK fork Replication origin Replicating Circular DNA Molecules Rolling Circle Replication Nucleotides are added to the Eongation of the Direction of 339 OH group displacing the 3 and continues rolling 539 Pterminated strand gt gt gt 339 0H A nuclease makes a cu yielding a 3 OH group iThe 5 Pterminated and a 539 P group strand also is copied DNA Replication is Very Fast E Coli completes replication of its 33 Mbp circular genome Has two growing points so rate is about 690 bpsecond Eur139 g1 r39I Human chromosome replication rate is ca 25 bp second To complete replication of its 29 Gbp genome in a reasonable time it has one origin every 90 kbp
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