Biotechnology

Biotechnology is an extended derivative of biochemistry & is a multidisciplinary field. It is an integrated subject of natural sciences and engineering sciences & achieves the application of organisms, cells, parts thereof and molecular analogues to produce products and services. It often refers to the production of products from raw materials with the aid of living organisms. The core principle of biotechnology involves harnessing biological systems and organisms, such as bacteria, yeast, and plants, to perform specific tasks or produce valuable substances. It has rendered a significant impact on many areas of society, from medicine to agriculture to environmental science.

 

One of the key techniques used in biotechnology is genetic-engineering, which allows scientists to modify the genetic makeup of organisms to achieve desired responses. This consists of inserting genes from one organism into another, creating new traits or modifying existing trait. Other important techniques used in biotechnology include tissue culture, which allows researchers to grow cells and tissues in the lab for research and medical purposes, and fermentation, which is used to produce a wide range of products such as beer, wine, and cheese.

 

The applications of biotechnology are very diverse and have led to the development of essential products like life-saving drugs, biofuels, genetically modified crops, and innovative materials. It has also been used to address environmental challenges, such as developing biodegradable plastics and using microorganisms to clean up contaminated sites. Therefore, biotechnology is a rapidly evolving field with significant potential to address pressing global challenges and improve the quality of life for people around the world; however, despite its numerous benefits, it also poses ethical and societal challenges, such as questions around genetic-modification and intellectual property rights.

 

Topics /Lesson Coverage:

FUNDAMENTALS OF MOLECULAR BIOTECHNOLOGY

 

I Fundamentals of Molecular Biotechnology 

 

1 The molecular biotechnology Revolution

Recombinant DNA Technology

Emergence of Molecular Biotechnology

Commercialization of Molecular Biotechnology

Concerns and consequences

 

2 Molecular Biotechnology Biological Systems

Prokaryotic and Eukaryotic organisms

Escherichia coli

Saccharomyces cerevisiae

Secretion Pathways in Prokaryotic and Eukaryotic Organisms

Eukaryotic Cells in Culture

 

3 DNA, RNA, and Protein Synthesis

Structure of DNA

DNA Replication

Decoding Genetic Information: RNA and protein Translation

Regulation of mRNA Transcription in Bacteria

Regulation of mRNA Transcription in Eukaryotes

 

4 Recombinant DNA Technology

Restriction Endonucleases

Plasmid Cloning Vectors

Plasmid Cloning Vector pBR322

Transformation and Selection

Other Plasmid Cloning Vector

Creating and Screening a Library

Making a Gene Library

Screening by DNA Hybridization

Screening by Immunological Assay

Screening by Protein Activity

Cloning DNA Sequences That Encode Eukaryotic Proteins

Vectors for Cloning Large Pieces of DNA

Bacteriophage λ Vectors

Cosmids

High-Capacity Bacterial Vector Systems

Genetic Transformation of Prokaryotes

Transferring DNA into E. Coli

Electroporation

Conjugation

 

 

5.   Chemical Synthesis, Sequencing, and Amplification of DNA

Chemical Synthesis of DNA

The Phosphonamidite Method

Uses of Synthesized Oligonucleotides

DNA Sequencing Techniques

Dideoxynucleotide Procedure for Sequencing DNA

Automated DNA Sequencing

Using Bacteriophage M13 as a DNA Sequencing Vector Primer

Walking

PCR

Gene Synthesis by PCR

Cycle Sequencing

 

6.Manipulation of Gene Expression in Prokaryotes

Gene Expression from strong and regulatable promoters

Regulatable Promoters

Increasing Protein Production

Large-Scale Systems

Expression in Other Microorganisms

Fusion Protein

Cleavages of Fusion Protein

Uses of Fusion Proteins

Surface Display

Unidirectional Tandem Gene Arrays

Translation Expression Vectors

Increasing Protein Stability

Protein Folding

Overcoming Oxygen Limitation

Use of Protease-Deficient Host Strains

Bacterial Haemoglobin

 

DNA Integration into the Host Chromosome

Removing Selectable Marker Genes

Increasing Secretion

L-Form Bacteria

Metabolic Load

 

7.   Heterologous Protein Production in Eukaryotic Cells

      Saccharomyces Cerevisiae Expression Systems

S. cerevisiae Vectors

Intracellular Production of Heterologous Proteins in S. Cerevisiae

Secretion of Heterologous Proteins by S. Cerevisiae

Pichia Pastoris and Other Yeast Expression Systems

Baculovirus-Insect Cell Expression Systems

Baculovirus Expression Vector System

Increasing the Yield of Recombinant Baculovirus

Construction of and E. Coil – Insect Cells Baculoviurs Shuttle Vector

Mammalian Glycosylation and Processing of Precursor Protiens in Insect Cells

Mammalian Cell Expression Systems

Selectable Marker Systems for Mammalian Expression Vectors

 

8.Directed Mutagenesis and Protein Engineering

Directed Mutagenesis Procedures

Oligonucleotide-Directed Mutagenesis with M13 DNA

Oligonucleotide-Directed Mutagenesis with Plasmid DNA

PCR-Amplified Oligonucleotide-Directed Mutagenesis

Random Mutagenesis with Degenerate Oligonucleotide primers

Random Mutagenesis with Nucleotide Analogues

Error-Prone PCR

DNA Shuffling

Mutant Proteins with Unusual Amino Acids

Protein Engineering

Adding Disulfide Bonds

Changing Asparagine to Other Amino Acids

Reducing the Number of Free Sulfhydryl Residues

Increasing Enzymatic Acvitity

Modifying Metal Cofactor Requirements

Decreasing Protease Sensitivity

Modifying Protein Specificity

Increasing Enzyme Stability and Specificity

Altering Multiple Properties

 

II Molecular biotechnology of Microbial Systems

 

9.     Molecular Diagnostics

Immunological Diagnostic Procedures

Enzyme-Linked Immunosorbent Assay

Monoclonal Antibodies

Formation and Selection of Hybrid Cells

Identification of Specific Antibody-Producing Hybrid Cells Lines

 

DNA Diagnostic Systems

Hybridization Probes

Diagnosis of Malaria

Detection of Trypanosoma

Nonradioactive Hybridization Procedures

Molecular Beacons

DNA Fingerprinting

Random Amplified Polymorphic DNA

Bacterial Biosensors

 

Molecular Diagnosis of Genetic Disease

Screening for Cystic Fibrosis

Sickle-Cell Anemia

The PCR/OLA procedure

Padlock Probes

Genotyping with Fluorescence-Labeled PCR Primers

 

10. Therapeutic Agents Pharmaceuticals

Isolation of Interferon cDNAs

Human Interferons

Human Growth Hormone

Tumor Necrosis Factor Alpha

Optimizing Gene Expression

Therapeutics Produced and Delivered by Intestinal Bacteria

 

Enzymes

DNase I

Alginate Lyase

Phenylalanine Ammoria Lyase

α1-Antitrypsin

 

Monoclonal Antibodies as Therapeutic Agents

Structure and Function of Antibodies

Preventing Rejection of Transplated Organs

Treating Brain Tumors

Chemically Linked Monoclonal Antibodies

Human Monoclonal Antibodies

Hybrid Human-Mouse Monoclonal Antibobies

Production of Antibodies in E. coli

Phase Combinatorial Libraries

Shuffling CDR Sequences

Single-Chain Antibodies

Nucleic Acids as Therapeutic Agents

Antisense RNA

Antisense Oligonucleotides

Ribozymes

Chimeric RNA-DNA Molecules

Interfering RNAs

Antibody Genes

Treating Genetic Disorders

Human Gene Therapy

Prodrug Activation Therapy

 

 

11. Vaccines

Subunit Vaccines

Herpes Simplex Virus

Foot-and-Mouth Disease

Peptide Vaccines

Genetic Immunization: DNA Vaccines

 

Attenuated Vaccines

Cholera

Salmonella Species

Leishmania Species

Herpes Simplex Virus

Vector Vaccines

Vaccines Directed against Viruses

Vaccines Directed against Bacteria

Bacteria as Antigen Delivery Systems

 

12. Synthesis of Commercial Products by RecombinantMicroorganisms

      Restriction Endonucleases

      Small Biological Molecules

      Synthesis of L-Ascorbic Acid

Microbial Syntheses of Indigo

Synthesis of Amino Acids

Removal of Lipids

Antibiotics

Cloning Antibiotic Biosynthesis Genes

Syntheses of Novel Antibiotics

Engineering Polyketide Antibiotics

Improving Antibiotic Production

Peptide Antibiotics

 

Biopolymers

Engineering Xanthomonas campestris for Xanthan Gum Production

Isolation of Melanin Biosynthesis Genes

Synthesis of an Animal Adhesive Biopolymers in Microbial Cells

Microbial Synthesis of Rubber

Microbial Production of Polyhydroxyalkanoates

 

13. Bioremediation and Biomass Utilization

      Microbial Degradation of Xenobiotics

Genetic Engineering of Biodegradation Pathways

Manipulation by Transfer of Plasmids

Manipulation by Gene Alteration

 

Utilization of Starch and Sugars

Commercial production of Fructose and Alcohol

Improving Alcohol Productions

Improving Fructose Production

Zymomonasmobilis

Silage Fermentation

 

Utilization of Cellulose

Components of Lignocellulose

Isolation of Prokaryotic Cellulase Genes

Isolation of Eukaryotic Cellulase Genes

Manipulation of Cellulase Genes

 

14  Plant Growth-Promoting Bacteria

      Nitrogen Fixation

Nitrogenase

Components of Nitrogenase

Genetic Engineering of the Nitrogenase Gene Cluster

Glycogen Synthase Mutants

Engineering Oxygen Levels

 

Hydrogenase

Hydrogen Metabolism

Genetic Engineering of Hydrogenase Genes

 

Nodulation

Competition among Nodulation Organisms

Genetic Engineering of Nodulation Genes

 

Growth Promotion by Free-Living Bacteria

Decreasing Plant Stress

 

Phytoremediation

Biocontrol of Pathogens

Siderophores

Antibiotics

Enzymes

Ice Nucleation and Antifreeze Proteins

Ethylene

Root Colonization

 

15. Microbial Insecticides

      Insecticidal Toxin of B. Thuringiensis

      Mode of Action and Use

Toxin Gene Isolation

Engineering of B. Thuringiensis Toxin Genes

Preventing the Development of Resistance

Genetic Engineering for Improve Biocontrol

Baculoviruses as Biocontrol Agents

Mode of Action

Genetic Engineering for Improved Biocontrol

 

16. Large-Scale Production of Protein from Recombinant Microorganisms

 

      Principle of Microbial Growth

      Batch Fermentation

Fed-Batch Fermentation

Continuous Fermentation

 

Maximizing the Efficiency of the Fermentation Process

High-Density Cell Cultures

Bioreactors

 

Typical Large-Scale Fermentation Systems

Two – Stage Fermentation in Tandem Airlift Reactors

Two-State Fermentation in a Single Stirred-Tank reactor

Batch versus Fed-Batch Fermentation

Harvesting Microbial Cells

Disrupting Microbial Cells

Downstream Processing

Protein Solubilization

Large-Scale Production of Plasmid DNA

 

III  Eukaryotic Systems

 

17. Genetic Engineering of Plants

 

Methodology

Plant Transformation with the Ti Plasmid of Agrobacterium tumefaciens

Ti Plasmid-Derived Vector Systems

Physical Methods of Transferring Genes to Plants

Microprojectile Bombardment

Use of Reporter Genes in Transformed Plant Cells

Manipulation of Gene Expression in Plants

Isolation and Use of Different Promoters

Targeted Alteration in plant DNA

Targeting Foreign DNA to the Chloroplast Genome

Secretion of Foreign Proteins

 

Production of Marker-Free Transgenic Plants

Removing Marker Genes from Nuclear DNA

Removing Marker Genes from Chloroplast DNA

 

18. Genetic Engineering of Plants: Applications

      Development of Insect-, Pathogen-, and Herbicide – Resistant plants

      Insect-Resistant Plants

Virus-Resistant Plants

Herbicide-Resistant Plants

Fungus-and Bacterium-Resistant Plants

 

Development of Stress-and Senescence-Tolerant Plants

Oxidative Stress

Salt Stress

Fruit Ripening and Flower Wilting

Genetic Manipulation of Flower Pigmentation

Modification of Plant Nutritional Content

      Amino Acids

      Lipids

      Vitamins

      Iron

 

      Modification of Food Plant and Appearance

      Preventing Discoloration

      Sweetness

      Starch

      Plants as Bioreactors

      Antibodies

      Polymers

      Foreign Proteins in Plants

      Edible Vaccines

      Plant Yield

      Increasing Iron Content

      Altering Lignin Content

      Increase Oxygen Content

 

19. Transgenic Animals

      Transgenic Mice: Methodology

      Retroviral Vector Method

DNA Microinjection Method

Engineered Embryonic Stem Cell Method

Genetic Modification with the Cre-loxP Recombination Systems

Transgenesis with High-Capacity Vectors

 

Transgenic Mice: Applications

Transgenic Models for Alzheimer Disease

Using Transgenic Mice as Test Systems

Conditional Regulation of Gene Expression

Conditional Control of Cell Death

Cloning Livestock by Nuclear Transfer

Transgenic Cattle, Sheep, Goats, and Pigs

Transgenic Birds

Transgenic Fish

 

20. Human Molecular Genetics

      Modes of Human Inheritance

Genetic Linkage and Gene Mapping

Detection and Estimation of Genetic Linkage in Humans

Genetic Mapping of Human Chromosomes

Genetic Polymorphism

Restriction Fragment Length Polymorphism

Short Tandem Repeat Polymorphism

 

Comprehensive Human Linkage Maps

Mapping of a Genetic Disease Locus to a Chromosome Location

Whole-Genome Sequence

 

Detection of Mutations in Human Genes

Single-Strand Conformation Analysis

Denaturing Gradient Gel Electrophoresis

Heteroduplex Analysis

Chemical Mismatch Cleavage

Direct DNA Sequencing

Determining Gene Function

 

IV    Regulating and Patenting Molecular

        Biotechnology

 

21    Regulating the Use of Biotechnology

        Regulating Recombinant DNA Technology

 

        Regulating Food and Food Ingredients  

        Chymosin

        Tryptophan

        Bovine Somatotropin

 

        Deliberate Release of GMOs

        Ice-Minus P. syringe   

        Open-Field Tests of Other GMOs  

        Controversy about GMOs  

 

        Human Gene Therapy   

        Development of a Policy for Somatic Cell Gene Therapy

        Accumulation of Defective Genes in Future Generations

        Human Germ Line Gene Therapy

        Cloning Human?        

 

22     Patenting Biotechnology Invention       

        Patenting

        Patenting in Different Countries   

        Patenting DNA Sequences  

Patenting Multicellular Organisms        

Patenting and Fundamental Research  

 

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