Lecture_Presentation_20 - Biotechnology

Chapter 20: DNA Tools and Biotechnology

• Lecture Presentations by: Nicole Tunbridge and Kathleen Fitzpatrick

• © 2021 Pearson Education, Inc.

Key Techniques in Biotechnology

Main Techniques

• DNA Sequencing: Determines the exact sequence of nucleotides in DNA.

• Gene Cloning: Involves replicating a specific gene to produce multiple copies.

• Expressing Genes: Producing gene products (proteins) in living systems.

• Analyzing Gene Expression: Understanding how genes are expressed in cells.

Applications

• Medicine: Development of treatments using cloned genes.

• Agriculture: Production of genetically modified crops for better yield.

• Environmental Cleanup: Using biotechnological methods for pollution management.

• Forensics: Applications in crime scene investigations and ancestry tracing.

DNA Technology

• DNA Technology: Techniques for sequencing and manipulating DNA.

• Nucleic Acid Hybridization: Base pairing between complementary DNA strands.

• Genetic Engineering: Direct manipulation of an organism's genes for practical benefits.

DNA Sequencing Techniques

First Generation - Sanger Sequencing

• Developed by Frederick Sanger, this method involves dideoxy or chain termination sequencing to determine a gene's nucleotide sequence.

Next-Generation Sequencing

• Includes rapid techniques that allow sequencing of many fragments simultaneously.

• Sequencing by Synthesis: Immobilizes DNA fragments and synthesizes complementary strands one nucleotide at a time.

Third-Generation Sequencing

• Even faster techniques that sequence long DNA molecules using a nanopore.

• Identification of bases based on interruptions in an electric current.

DNA Cloning

• DNA Cloning: Producing multiple identical copies of DNA segments using plasmids.

• Plasmids: Small circular DNA molecules that replicate independently in bacterial cells.

Process of Cloning

1. Insertion of DNA: DNA from one source is inserted into a plasmid to create a recombinant DNA molecule.

2. Transformation: Plasmid is introduced into a bacterial cell.

3. Cell Growth: Bacteria are cultured to produce clones containing the gene of interest.

Gene Cloning Applications

• Human growth hormone and blood clotting proteins produced for medical use.

Techniques for Creating Recombinant DNA

Using Restriction Enzymes

• Restriction Enzymes: Cut DNA at specific sequences (restriction sites).

• Most useful enzymes create sticky ends to facilitate the joining of DNA fragments.

DNA Ligase

• An enzyme that joins the fragments of DNA by sealing the bonds between restriction fragments, forming recombinant DNA.

Gel Electrophoresis

• Used to separate DNA fragments for visualization and verification of recombinant plasmids.

Polymerase Chain Reaction (PCR)

Overview

• PCR amplifies specific DNA segments through a series of temperature cycles which include:

  • Denaturation: Heating to separate strands.

  • Annealing: Cooling to allow primers to bind to target sequences.

  • Extension: DNA polymerase synthesizes new DNA strands.

Taq Polymerase

• A heat-stable DNA polymerase critical for PCR's effectiveness.

Applications of PCR

• Amplification from ancient DNA sources, forensic samples, and other low-quantity biological samples.

Conclusion

• Cloned genes can be expressed in bacteria or eukaryotic cells for research or practical applications.

• The techniques discussed play a vital role in advancing biotechnology, genetics, and molecular biology.