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First generation Gene Sequencing

First generation Gene Sequencing. Jessie Marlenee BIOL 546. https://en.wikipedia.org/wiki/Phi_X_174. Recap. Watson & Crick discover DNA double helix 1953 Restriction Enzymes Southern Blotting Cloning. https://en.wikipedia.org/wiki/Nucleic_acid_double_helix.

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First generation Gene Sequencing

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  1. First generation Gene Sequencing Jessie Marlenee BIOL 546 https://en.wikipedia.org/wiki/Phi_X_174

  2. Recap • Watson & Crick discover DNA double helix 1953 • Restriction Enzymes • Southern Blotting • Cloning https://en.wikipedia.org/wiki/Nucleic_acid_double_helix https://en.wikipedia.org/wiki/Restriction_enzyme

  3. Initial sequencing efforts • Proteins • RNA • Holley and colleagues (1965) • alanine tRNA from Saccharomyces cerevisiae • Wu and Kaiser (1968-71) • Incorporation of radiolabeled nucleotides into strand to determine sequence • Walter Fiers' laboratory (1972-76) • Coat protein of bacteriophage MS2, followed by complete genome four years later (3569 nucleotides of ssRNA) • Replacement of 2D fractionation https://en.wikipedia.org/wiki/Bacteriophage_MS2 Kaiser, A. D. and Wu, R. 1968.

  4. Sanger’s “Plus and Minus” technique(1975) https://en.wikipedia.org/wiki/Phi_X_174 https://slideplayer.com/slide/5188821/

  5. Maxam and Gilbert’s Chemical cleavage technique (1977) Matthews and van Holde: Biochemistry, 2nd edition

  6. Sanger’s “chain termination” technique (1977) Also known as the dideoxynucleotide method or sanger sequencing

  7. 1980 Nobel prize in chemistry • ½ Awarded to Paul Berg • ¼ Awarded to Walter Gilbert • ¼ Awarded to Frederick Sanger

  8. References • Franca, L. T., Carrilho, E., and Kist, T. B. 2002. A review of DNA sequencing techniques. Q. Rev. Biophys. 35:169–200. • Guzvic, M. 2013. The history of DNA sequencing. J Med Biochem. 32:301–12. • Heather, J. M., and Chain, B. 2016. The sequence of sequencers: The history of sequencing DNA, Genomics. 107:1-8. • Hutchison C. A. III. 2007. DNA sequencing: bench to bedside and beyond. Nucleic Acids Res. 35:6227–6237. • Holley, R. W., Apgar, J., Everett, G. A., Madison, J. T., Marquisee, M., Merrill, S. H., Penswick, J. R. and Zamir, A. 1965. Structure of a ribonucleic acid. Science. 147:1462–1465. • Kaiser, A. D. and Wu, R. 1968. Structure and function of DNA cohesive ends. Cold Spring Harb. Symp. Quant. Biol. 33:729–734. • Mardis, E. R. 2013. Next-generation sequencing platforms. Annu. Rev. Anal. Chem. 6:287–303. • Maxam, A. M. & Gilbert, W. 1977. A new method for sequencing DNA. Proc. Natn. Acad. Sci. 74:560–564. • Sanger, F. and Coulson, A. R. 1975. A rapid method for determining sequences in DNA by primed synthesis with DNA polymerase. J. Mol. Biol., 94:441–448. • Sanger, F., Nicklen, S. & Coulson, A. R. 1977. DNA sequencing with chain-terminating inhibitors. Proc. Natn. Acad. Sci. 74:5463–5467. • The Nobel Prize in Chemistry 1980. NobelPrize.org. Nobel Media AB 2018. Thu. 13 Sep 2018. https://www.nobelprize.org/prizes/chemistry/1980/summary/ • Wu, R. and Kaiser, A. D. 1968. Structure and base sequence in the cohesive ends of bacteriophage lambda DNA. J. Mol. Biol., 35:523–537.

  9. Questions?

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