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.