SM

Intro Genetic Engineering Flashcards

Introduction to Genetic Engineering

Didactic Unit 1: Theoretical Foundations

  • Introduction to Genetic Engineering.
  • Basic lab techniques.
  • Basic enzymology for in vitro DNA manipulation.
  • Molecular Cloning I: Basic Vectors.
  • Molecular Cloning II: Cellular cloning systems.
  • Molecular Cloning III: Cloning Methods.
  • Advanced vectors in bacteria and other organisms.
  • Gene Libraries.
  • Expression of cloned genes.
  • Modification of DNA sequences: mutagenesis and gene editing.
  • Synthetic Biology.

A History of Molecular Cloning

  • 1952: Genetic demonstration of phage restriction.
  • 1953: Genetic recombination demonstrated.
  • 1967: Isolation of the first DNA ligases.
  • 1968: Isolation of the first restriction factor.
  • 1970: Isolation of restriction enzymes (RES).
  • 1971: Restriction enzyme mapping of the simian virus SV40.
  • 1972: Assembly of the first recombinant DNA & transformation of the first E.coli.
  • 1974: New England Biolabs® opens for business.
  • 1975: Introduction of Maxam-Gilbert and Sanger sequencing.
  • 1976: Launch of REBASE® (Restriction Enzyme Database).
  • 1977: Report of the first cloning vector (pBR322).
  • 1978: Nobel Prize awarded to Smith, Arber and Nathans for the discovery of RES.
  • 1983: Introduction of PCR.
  • 1985: Generation of pUC plasmids.
  • 1993: Mullis shares the Nobel prize for his work on PCR.
  • 2000: In vitro synthesis of a whole genome.
  • 2003: Development of first standardized DNA parts system, BioBricks, by Knight.
  • 2008: Publication of the original Golden Gate Assembly paper.
  • 2010: First synthetic life- Mycoplasma mycoides.
  • 2011: NEB publishes rules and tools for Data-optimized Design of Golden Gate Assembly (GGA).
  • 2014: First complete eukaryotic chromosome synthesized (yeast).
  • 2018: NEB publishes 40 kb phage genome construction.
  • 2019: Chin publishes development of Syn61 E. coli strain with fully recoded genome.
  • 2022: Development of CRISPR-Cas9 gene editing technology.

Eras of Genomics

  • Pre-genomic era: Restriction enzymes discovered, rDNA molecules generated, Rapid DNA sequencing developed, First viral genome sequenced.
  • Genomic era: First bacterial genome sequenced, Human Genome Project started, Human genome completed , Second-generation sequencing technology.
  • Post-genomic era: Third-generation sequencing technology, Genome editing, Systems and synthetic biology, First trial of gene editing in vivo, T2T human genome sequence.

Course Big Questions

  • How can I use Genetic Engineering to make a better world?
  • Means learning how to perform Molecular Cloning and what to perform Molecular Cloning for.
  • The four steps in DNA cloning:
    • Generation of DNA fragments: Source of DNA (In vitro).
    • Joining to vector.
    • Introduction into host cell (In vivo).
    • Selection of cloned sequence. Further processing.

Cloning Strategies

  • Cloning strategies are influenced by whether the genome sequence is known or not. If not, consider sequencing the genome using NGS.
  • If the genome sequence is known, use a bioinformatics approach (in silico).
  • Objective defines the cloning strategy.