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DNA. Searching for Genetic Material. Mendel: modes of heredity in pea plants (1850’s) Morgan: genes located on chromosomes (early 1900’s) Griffith: bacterial work (1920’s) transformation - change in genotype and phenotype due to assimilation of external substance (DNA) by a cell
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Searching for Genetic Material • Mendel: modes of heredity in pea plants (1850’s) • Morgan: genes located on chromosomes (early 1900’s) • Griffith: bacterial work (1920’s) • transformation- change in genotype and phenotype due to assimilation of external substance (DNA) by a cell • Avery: transformation agent was DNA (1944)
Hershey-Chase Experiment • 1952 Experiment with bacteriophages • viruses that infect bacteria • Tested if DNA or protein was the hereditary material • in T2 (a bacteriophage that infects E. coli) • Sulfur (S) is in protein, phosphorus (P) is in DNA • Only P was found in host cell
DNA Structure • Chargaff • ratio of nucleotide bases (A=T; C=G) • Structure of DNA researched by Pauling and Wilkins/Franklin • Watson & Crick (1953) • Proposed the structure of DNA after viewing an X-ray diffraction photo by Rosalind Franklin
The Double Helix • Nucleotides: nitrogenous base (thymine, adenine, cytosine, guanine); sugar (deoxyribose); phosphate group
Base Pairing Rules • Purines: A & G • Pyrimidines: C & T (Chargaff rules) • A’s H+ bonds (2x) with T and C’s H+ bonds (3x) with G • Van der Waals attractions between the stacked pairs
DNA Replication • Watson & Crick strands are complementary; nucleotides line up on template according to base pair rules • Meselson & Stahlreplication is semiconservative; Expt: varying densities of radioactive nitrogen
3 Replication Models • Meselson and Stahl concluded that DNA follows the semiconservative model
DNA Replication in Action • Origin of replication (“bubbles”)= beginning of replication • Replication fork: Y-shaped region where new strands of DNA are elongating • Helicase: catalyzes the untwisting of the DNA at the replication fork • DNA polymerase: catalyzes the elongation of new DNA
Antiparallel Nature • Sugar/phosphate backbone runs in opposite directions (Crick) • One strand runs 5’ to 3’, while the other runs 3’ to 5’ • DNA polymerase only adds nucleotides at the free 3’ end, forming new DNA strands in the 5’ to 3’ direction only
Leading and Lagging • Leading strand: • synthesis toward the replication fork (only in a 5’ to 3’ direction from the 3’ to 5’ master strand) • Lagging strand: • synthesis away from the replication fork in small pieces (Okazaki fragments) • joined by DNA ligase
The Lagging Strand • Primase enzyme attaches a primer • a short sequence of RNA (sometimes DNA) • Small segments replicated 5’ 3’ • After strand replicates, primer falls off
Proofreading • Mismatch repair: DNA polymerase • Excision repair: nuclease
Shortening Ends • Because lagging strands need 3’ end, replication cannot extend to the very end of a DNA strand • Chromosomes contain telomeres • Repeating sequences of DNA • Don’t contain genes • Telomerase lengthens telomeres so that they don’t continually get shorter • Chromosomes can divide without losing genes