Presentation on theme: "Mark Mayo Cypress College"— Presentation transcript:
1Mark Mayo Cypress College The History of DNAMark MayoCypress CollegeLast update 9/16/13
2Transformation Frederick Griffith (1923) Used healthy miceMice were injected with either R(rough) strain of Streptococcus pneumoniae. The mice live and their immune system kills R bacteria. No live bacteriaMice injected with the S (smooth) strain of Streptococcus pneumoniae. The mice die. The dead mice have live S bacteria.Mice injected with heat-killed S strain. Mice live with no live bacteria found in mice.Mice injected with mixture of live R strain and heat-killed S strain. Mice die and live S strain bacteria are found in the dead mice.Heat does not destroy the active factor that is responsible for heredity (DNA).It is said that the bacteria were transformed by this active agent.
4Transformation II Avery, McCarty, Macleod (1944) Think re-mix of GriffithRepeated Griffith’s work, but knew that DNA was the substance of transformationSeparated classes molecules from s cell debrisTested each fraction for transforming ability, one at a timeOnly DNA transformed r cells into s cellsTo provide r cells with s DNA is to provide r cells with s genes
6DNA or Protein as Active Agent Alfred Hershey and Martha Chase DNA or Protein as Active Agent Alfred Hershey and Martha Chase* (1950’s)Used radioactive labels on bacteriophage components to decide if DNA or protein was the transforming factorLabel viral protein with S35 *Label viral DNA with P32 *Allow infectionWash viral particles (with blender!)Check for label after subsequent infection into new bacteriaFound only P32Hence DNA is the transforming factor
7DNA or Protein as Active Agent Alfred Hershey and Martha Chase
8Chargaff’s Rules Erwin Chargaff (1949) He studied the relative amounts of each nucleic acid base in a great variety of plant and animal speciesRoughly found that A=T and G=C, but not exactly due to errors in the technology!Purines are exactly equal to pyrimidinesHis methodology for the time was good, but now we get exact amountsHe could not make the connection (Watson and Crick used his data however)
10Alpha Helix Linus Pauling (1948-1950) Worked with proteins and determined that collagen has a helical arrangement for its polypeptidesHe called the helix an alpha helixHe suspected that DNA might also have a helical arrangement, but could not get it to compute with a single strandPauling suggested DNA had a triple helix, but had no proofWatson and Crick heard his idea about a helix…
12X Ray Diffraction Maurice Wilkins and Rosalyn Franklin Used Xray diffraction to study proteins and other moleculesDNA was very difficult to crystallize and a tough candidate for Xray diffractionRosalyn Franklin was a very talented graduate student in the lab of WilkinsShe was successful at crystallizing DNA in two forms A and BThe forms on an X was seen indicating some kind of helixShe could measure the distances between repeating units on the moleculeShe could also measure the diameter of the moleculeWilkins sent her unpublished data to Watson and Crick without her permissionShe died before the Nobel prize or she might have shared it
13X Ray Diffraction Maurice Wilkins and Rosalyn Franklin
14Semi-conservative DNA Replication Matthew Meselsohn and Frank Stahl They used two isotopes of Nitrogen – 14N and 15N15N is heavier than 14NThey grew bacteria for several generations in heavy 15N (all DNA would be heavy!)Abruptly changed the medium to lighter 14N for one or two generationsUsed density-gradient ultracentrifugation to separate the DNA strands by weightAfter one generation all DNA was medium between heavy and light (thus SEMI-CONSERVATIVE)After two generation DNA had two bands: medium and light.
15Semi-conservative DNA Replication Matthew Meselsohn and Frank Stahl This one would be one heavy and one light bandHalf light and half heavy = medium weight
16Discontinuous DNA Replication Reiji Okazaki He knew about DNA polymeraseIt moves from 5’ to 3’ only on the leading strandHe searched for a second polymerase that worked in the reverse directionAfter unsuccessfully searching he used his brillianceFound numerous small fragments and also long segments as DNA was replicatedFinally decided that the short segments were from the lagging strand *DNA polymerase worked on both sides continuously on the leading strand and in several places on the lagging strandDNA ligase connects the small portions (now called Okazaki fragments) * on the lagging strand