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Transcript
DNA and Replication 1 History of DNA 2 History of DNA • Early scientists thought protein was the cell’s hereditary material because it was more complex than DNA • Proteins were composed of 20 different amino acids in long polypeptide chains 3 Transformation • Fred Griffith worked with virulent S and nonvirulent R strain Pneumoccocus bacteria • He found that R strain could become virulent when it took in DNA from heat-killed S strain • Study suggested that DNA was probably the genetic material 4 Griffith Experiment 5 History of DNA • Chromosomes are made of both DNA and protein • Experiments on bacteriophage viruses by Hershey & Chase proved that DNA was the cell’s genetic material Radioactive 32P was injected into bacteria! 6 Discovery of DNA Structure • Erwin Chargraff showed the amounts of the four bases on DNA ( A,T,C,G) • In a body or somatic cell: A = 30.3% T = 30.3% G = 19.5% C = 19.9% 7 Chargaff’s Rule • Adenine must pair with Thymine • Guanine must pair with Cytosine • The bases form weak hydrogen bonds T A G C 8 DNA Structure • Rosalind Franklin took diffraction x-ray photographs of DNA crystals • In the 1950’s, Watson & Crick built the first model of DNA using Franklin’s x-rays 9 Rosalind Franklin 10 DNA Structure 11 DNA • Two strands coiled called a double helix • Sides made of a pentose sugar Deoxyribose bonded to phosphate (PO4) groups by phosphodiester bonds • Center made of nitrogen bases bonded together by weak hydrogen bonds 12 DNA Double Helix “Rungs of ladder” Nitrogenous Base (A,T,G or C) “Legs of ladder” Phosphate & Sugar Backbone 13 Helix • Most DNA has a right-hand twist with 10 base pairs in a complete turn • Left twisted DNA is called Z-DNA or southpaw DNA • Hot spots occur where right and left twisted DNA meet producing mutations 14 DNA • Stands for Deoxyribonucleic acid • Made up of subunits called nucleotides • Nucleotide made of: 1. Phosphate group 2. 5-carbon sugar 3. Nitrogenous base 15 DNA Nucleotide Phosphate Group O O=P-O O 5 CH2 O N C1 C4 Sugar (deoxyribose) C3 C2 Nitrogenous base (A, G, C, or T) 16 Pentose Sugar • Sugars are numbered clockwise 1’ to 5’ 5 CH2 O C1 C4 Sugar (deoxyribose) C3 C2 17 5 DNA O 3 3 P 5 O O C G 1 P 5 3 2 4 4 P 5 P 2 3 1 O T A 3 O 3 5 O 5 P P 18 Antiparallel Strands • One strand of DNA goes from 5’ to 3’ (sugars) • The other strand is opposite in direction going 3’ to 5’ (sugars) 19 Nitrogenous Bases • Double ring PURINES Adenine (A) Guanine (G) A or G • Single ring PYRIMIDINES Thymine (T) Cytosine (C) T or C 20 Base-Pairings • Purines only pair with Pyrimidines • Three hydrogen bonds required to bond Guanine & Cytosine 3 H-bonds G C 21 •Two hydrogen bonds are required to bond Adenine & Thymine T A 22 Question: • If there is 30% Adenine, Adenine how much Cytosine is present? 23 Answer: • There would be 20% Cytosine • Adenine (30%) = Thymine (30%) • Guanine (20%) = Cytosine (20%) • Therefore, 60% A-T and 40% C-G 24 DNA Replication 25 Replication of DNA • DNA is the only molecule that can replicate itself. This is accomplished through a process known as replication. 26 • They attach themselves at their complementary bases. • A series of enzymes called polymearases fuse the free nucleotides together in the complimentary chain of DNA. • The free floating nucleotides in your cells are derived from the food you eat. Steak supplies you with muscle cells from a cow. Does not mean you will turn into a cow. Specialized enzymes in your digestive tract break down the cow DNA into cucleotides which you use to make human DNA 27 • Genetic mistakes do occur but are very infrequent • Environmental factors such as hazardous chemicals or radiation are one source of genetic mistakes that do occur. • Can cause uncomplimentary bases to join. • Permanent damage is prevented by enzyme that act as proof readers. They run along the strands of DNA looking for mismatched pairs. It snips the error and replaces it with the correct nucleotide. 28 Replication • the weak hydrogen bonds that hold the complementary nitrogen bases together are broken. The two edges of the ladder seem to unzip. • The parent strands are conserved (remain intact) • This is called semiconservative replication • semiconservative replication produces two “half old, half new” strands of DNA. • Each parent strand acts as a template or mold to which free floating nucleotides in29 the cell can attach. • LAB page 525 required - photocopy 30 Lab 31 Replication Facts • DNA has to be copied before a cell divides • DNA is copied during the S or synthesis phase of interphase • New cells will need identical DNA strands 32 Synthesis Phase (S phase) • S phase during interphase of the cell cycle • Nucleus of eukaryotes DNA replication takes place in the S phase. S phase G1 interphase G2 Mitosis -prophase -metaphase -anaphase -telophase 33 DNA Replication • Begins at Origins of Replication • Two strands open forming Replication Forks (Y-shaped region) • New strands grow at the forks 5’ Parental DNA Molecule 3’ 3’ Replication Fork 34 5’ DNA Replication • As the 2 DNA strands open at the origin, Replication Bubbles form • Eukaryotic chromosomes have MANY bubbles • Prokaryotes (bacteria) have a single bubble Bubbles Bubbles 35 DNA Replication • Enzyme Helicase unwinds and separates the 2 DNA strands by breaking the weak hydrogen bonds • Single-Strand Binding Proteins attach and keep the 2 DNA strands separated and untwisted 36 DNA Replication • Enzyme Topoisomerase attaches to the 2 forks of the bubble to relieve stress on the DNA molecule as it separates Enzyme Enzyme DNA 37 DNA Replication • • • Before new DNA strands can form, there must be RNA primers present to start the addition of new nucleotides Primase is the enzyme that synthesizes the RNA Primer DNA polymerase can then add the new nucleotides 38 39 DNA Replication • DNA polymerase can only add nucleotides to the 3’ end of the DNA • This causes the NEW strand to be built in a 5’ to 3’ direction 5’ 3’ Nucleotide DNA Polymerase Direction of Replication RNA Primer 40 5’ Remember HOW the Carbons Are Numbered! Phosphate Group O O=P-O O 5 CH2 O N C1 C4 Sugar (deoxyribose) C3 C2 Nitrogenous base (A, G, C, or T) 41 Remember the Strands are Antiparallel 5 O 3 3 P 5 O O C G 1 P 5 3 2 4 4 P 5 P 2 3 1 O T A 3 O 3 5 O 5 P P 42 Synthesis of the New DNA Strands • The Leading Strand is synthesized as a single strand from the point of origin toward the opening replication fork 5’ 3’ Nucleotides DNA Polymerase 5’ RNA Primer 43 Synthesis of the New DNA Strands • The Lagging Strand is synthesized discontinuously against overall direction of replication • This strand is made in MANY short segments It is replicated from the replication fork toward the origin Leading Strand 5’ 3’ DNA Polymerase 5’ 3’ Lagging Strand RNA Primer 3’ 5’ 3’ 5’ 44 Lagging Strand Segments • Okazaki Fragments - series of short segments on the lagging strand • Must be joined together by an enzyme DNA Okazaki Fragment RNA Primer 5’ 3’ Polymerase Lagging Strand 3’ 5’ 45 Joining of Okazaki Fragments • The enzyme Ligase joins the Okazaki fragments together to make one strand DNA ligase 5’ 3’ Okazaki Fragment 1 Okazaki Fragment 2 3’ 5’ Lagging Strand 46 Replication of Strands Replication Fork Point of Origin 47 Proofreading New DNA • DNA polymerase initially makes about 1 in 10,000 base pairing errors • Enzymes proofread and correct these mistakes • The new error rate for DNA that has been proofread is 1 in 1 billion base pairing errors 48 Semiconservative Model of Replication • Idea presented by Watson & Crick • The two strands of the parental molecule separate, and each acts as a template for a new complementary strand • New DNA consists of 1 PARENTAL (original) and 1 NEW DNA Template strand of DNA Parental DNA New DNA 49 DNA Damage & Repair • Chemicals & ultraviolet radiation damage the DNA in our body cells • Cells must continuously repair DAMAGED DNA • Excision repair occurs when any of over 50 repair enzymes remove damaged parts of DNA • DNA polymerase and DNA ligase replace and bond the new nucleotides together 50 Question: • What would be the complementary DNA strand for the following DNA sequence? DNA 5’-CGTATG-3’ 51 Answer: DNA DNA 5’-GCGTATG-3’ 3’-CGCATAC-5’ 52 53