Slide 1

Biotechnology: Principles and Processes

• Introduction to biotechnology
• Genetic engineering
• Applications of biotechnology
• Competent cells

Slide 2

Introduction to Biotechnology

• Definition: The use of living organisms or their products to modify or improve human health, agriculture, and the environment
• Historical background: Biotechnology has been used for centuries in processes such as brewing, bread-making, and cheese production
• Modern biotechnology: Involves manipulating genes and DNA to develop new products and processes

Slide 3

Genetic Engineering

• Definition: The process of manipulating an organism’s DNA to achieve desired traits or characteristics
• Techniques used in genetic engineering:
  • Recombinant DNA technology: Combining DNA from different sources to create novel sequences
  • Polymerase Chain Reaction (PCR): Amplifying specific DNA sequences for analysis
  • DNA sequencing: Determining the order of nucleotides in a DNA molecule
  • Gene cloning: Creating multiple copies of a specific gene
  • Genetic transformation: Introducing foreign DNA into an organism

Slide 4

Applications of Biotechnology

• Medicine and healthcare:
  • Production of pharmaceuticals, such as insulin and vaccines
  • Gene therapy: Correcting genetic disorders by replacing faulty genes
  • Diagnosis of diseases through DNA testing

Slide 5

• Agriculture:
  • Genetically modified crops for increased yield, pest resistance, and herbicide tolerance
  • Improved livestock breeds through genetic selection and cloning

Slide 6

• Environmental applications:
  • Bioremediation: The use of organisms to degrade pollutants in the environment
  • Bioplastics: Development of environmentally-friendly plastics using biodegradable polymers
  • Biofuels: Production of renewable energy sources, such as biodiesel and bioethanol

Slide 7

Competent Cells

• Definition: Cells that have been modified to uptake and express foreign DNA
• Process of making competent cells:
  • Cells are treated with chemicals or subjected to physical methods to increase their permeability
  • Foreign DNA is introduced into the cells through transformation
  • Successful transformation results in the expression of the foreign DNA

Slide 8

• Applications of competent cells:
  • Cloning: Competent cells are often used to create multiple copies of a specific gene
  • Protein expression: Competent cells can be used to produce recombinant proteins for various applications
  • Genetic engineering experiments: Competent cells provide a platform for manipulating and studying DNA

Slide 9

Summary

• Biotechnology is the use of living organisms or their products to modify or improve human health, agriculture, and the environment
• Genetic engineering involves manipulating an organism’s DNA to achieve desired traits or characteristics
• Applications of biotechnology include medicine, agriculture, and environmental applications
• Competent cells are modified cells that can uptake and express foreign DNA

Slide 10

Summary (contd.)

• Making competent cells involves increasing their permeability and introducing foreign DNA through transformation
• Competent cells find applications in cloning, protein expression, and genetic engineering experiments
• Biotechnology offers immense potential in various fields and continues to play a significant role in scientific advancements.

Slide 11

Biotechnology: Principles and Processes - Competent Cells

• Properties of competent cells:

  • High transformation efficiency
  • Ability to incorporate foreign DNA into their genome
  • Ability to express the foreign DNA

• Types of competent cells:

  • Chemically competent cells
  • Electrocompetent cells

• Factors affecting competency:

  • Growth phase of cells
  • Temperature and duration of treatment
  • Composition of transformation medium

Slide 12

Chemically Competent Cells

• Preparation of chemically competent cells:

  • Cells are grown to mid-log phase
  • Treated with a competent cell preparation buffer
  • Subjected to a heat shock or cold shock
  • Incubated in recovery medium
  • Spread on selective agar plates

• Example: Escherichia coli (E. coli) chemically competent cells

Slide 13

Electrocompetent Cells

• Preparation of electrocompetent cells:

  • Cells cultured to mid-log phase
  • Washed and resuspended in an ice-cold electroporation buffer
  • Electroporated using an electrical pulse
  • Incubated in recovery medium
  • Spread on selective agar plates

• Example: Saccharomyces cerevisiae (yeast) electrocompetent cells

Slide 14

Factors Affecting Competency

• Growth phase of cells:

  • Cells in mid-log phase have higher transformation efficiency
  • Optimal time for transformation varies depending on the organism

• Temperature and duration of treatment:

  • Heat or cold shock can increase the permeability of the cell membrane
  • Optimal temperature and duration may differ for different organisms

Slide 15

Factors Affecting Competency (contd.)

• Composition of transformation medium:

  • Presence of cations (Ca2+ and Mg2+) can improve the competency of cells
  • Use of specific salts, such as calcium chloride, can enhance transformation efficiency

• Other factors:

  • DNA concentration and quality
  • Cell density
  • Recovery conditions after transformation

Slide 16

Examples of Competent Cell Applications

• Cloning:

  • Transforming competent cells with a plasmid containing the gene of interest
  • Allows for the amplification and propagation of the desired DNA fragment

• Protein expression:

  • Transformation of competent cells with a plasmid carrying the gene encoding the protein of interest
  • Cells will produce and express the recombinant protein

Slide 17

Examples of Competent Cell Applications (contd.)

• Gene editing:

  • Introduction of CRISPR-Cas9 or other gene editing systems into competent cells
  • Enables targeted modifications of the genome

• Biotechnology research:

  • Transformation of competent cells with various constructs for studying gene function, regulation, and interactions

Slide 18

Summary

• Competent cells possess the ability to uptake and express foreign DNA
• Chemically competent cells and electrocompetent cells are the two main types
• Factors affecting competency include growth phase, treatment temperature and duration, and transformation medium composition
• Examples of competent cell applications include cloning, protein expression, gene editing, and biotechnology research

Slide 19

Summary (contd.)

• Competent cells play a crucial role in various biotechnological processes
• Understanding the properties and preparation methods of competent cells is essential in modern biotechnology research and applications

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Slide 21

Gene Cloning

• Definition: The process of creating multiple copies of a specific gene
• Steps involved in gene cloning:
  • Isolation of the gene of interest
  • Insertion of the gene into a cloning vector, such as a plasmid
  • Transformation of competent cells with the recombinant plasmid
  • Selection and identification of transformed cells

Slide 22

Gene Cloning (contd.)

• Applications of gene cloning:

  • Production of recombinant proteins for research and therapeutic purposes
  • Study of gene function through gene knockout and overexpression experiments
  • Creation of genetically modified organisms with desired traits

• Example: Cloning the gene for human insulin and producing it in E. coli bacteria

Slide 23

Polymerase Chain Reaction (PCR)

• Definition: A laboratory method used to amplify specific DNA sequences
• Steps involved in PCR:
  • Denaturation: Heating the DNA to separate the double-stranded template into single strands
  • Annealing: Cooling the reaction to allow primers to bind to the complementary DNA strands
  • Extension: Raising the temperature to activate DNA polymerase, which extends the primers and synthesizes new DNA strands

Slide 24

Polymerase Chain Reaction (PCR) (contd.)

• Applications of PCR:

  • DNA sequencing
  • Gene cloning
  • Diagnosis of genetic disorders
  • Forensic DNA analysis

• Example: Using PCR to amplify a DNA fragment for analysis or cloning

Slide 25

DNA Sequencing

• Definition: Determining the order of nucleotides in a DNA molecule
• Techniques used in DNA sequencing:
  • Sanger sequencing: A chain-termination method using dideoxynucleotides
  • Next-generation sequencing (NGS): High-throughput sequencing methods, such as Illumina sequencing and Ion Torrent sequencing

Slide 26

DNA Sequencing (contd.)

• Applications of DNA sequencing:

  • Genome sequencing: Determining the complete DNA sequence of an organism’s genome
  • Identification of genetic variations and mutations
  • Comparative genomics and evolutionary studies

• Example: Sequencing the human genome as part of the Human Genome Project

Slide 27

Gene Therapy

• Definition: Correcting genetic disorders by replacing faulty genes or introducing functional genes into cells
• Approaches used in gene therapy:
  • Gene addition: Introducing a functional gene into the patient’s cells
  • Gene editing: Modifying the existing gene sequence using techniques like CRISPR-Cas9

Slide 28

Gene Therapy (contd.)

• Applications of gene therapy:

  • Treatment of genetic disorders, such as cystic fibrosis and muscular dystrophy
  • Cancer therapy: Targeting tumor cells with modified genes to inhibit their growth
  • Immunotherapy: Modifying immune cells to enhance their anti-cancer activity

• Example: Using gene therapy to cure severe combined immunodeficiency (SCID)

Slide 29

Bioremediation

• Definition: The use of living organisms to degrade or eliminate environmental pollutants
• Types of bioremediation:
  • In situ bioremediation: Treating pollutants within their natural environment
  • Ex situ bioremediation: Removing contaminated material for treatment in a controlled environment

Slide 30

Bioremediation (contd.)

• Applications of bioremediation:

  • Cleaning up oil spills and contaminated groundwater
  • Removing heavy metals and hazardous chemicals from soil and water
  • Restoring ecosystems affected by pollution

• Example: Using bacteria to degrade oil spills in marine environments

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