A low-cost approach to PCR : appropriate transfer of biomolecular techniques /

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The polymerase chain reaction (PCR) is a technique used to replicate specific pieces of DNA millions of times, which permits the detection and analysis of minute amounts of nucleic acids. Since its introduction in the late 1980s, this technique has been applied not only in molecular biology research...

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Detalles Bibliográficos
Clasificación:Libro Electrónico
Autor principal: Harris, Eva, 1965-
Otros Autores: Kadir, Nazreen
Formato: Electrónico eBook
Idioma:Inglés
Publicado: New York : Oxford University Press, 1998.
Temas:
Molecular biology > Technology transfer.
Polymerase chain reaction > Laboratory manuals.
Polymerase chain reaction.
Polymerase Chain Reaction
Molecular Biology
Réaction en chaîne de la polymérase > Manuels de laboratoire.
Réaction en chaîne de la polymérase.
SCIENCE > Life Sciences > Molecular Biology.
Biologia molecular.
Reaccion en cadena de la polimerasa.
Transferencia de tecnologia.
Methode
Polymerase-Kettenreaktion
Vereinfachung
Laboratory Manual
Laboratory manuals.
Manuels de laboratoire.
Acceso en línea:Texto completo
Tabla de Contenidos:
  • 1 Introduction
  • 2 PCR Technology
  • 2.1. Description of the Technique
  • 2.1.1. Historical Overview
  • 2.1.2. The Molecular Basis of PCR
  • 2.1.3. Advantages and Disadvantages
  • 2.2. Technical Details
  • 2.2.1. Components of the PCR Mixture
  • 2.2.2. Thermal Cycling Parameters
  • 2.2.3. Optimizing
  • 2.2.4. Potential Problems
  • 2.2.5. Detection of Products
  • 2.3. Frequently-Used PCR-Based Techniques
  • 3 Principles of Sustainable Technology Transfer
  • 3.1. A Low-Cost Methodology
  • 3.1.1. Appropriate Technology
  • 3.1.2. Alternative Techniques
  • 3.1.3. Simplification of Protocols
  • 3.1.4. In-House Preparation of Reagents
  • 3.1.5. Recycling
  • 3.1.6. Donated Materials
  • 3.2. Knowledge-Based Participatory Transfer Process
  • 3.3. Appropriate Application
  • 3.3.1. General Considerations
  • 3.3.2. An Evaluation Framework
  • 3.3.3. Assessment Criteria for PCR
  • 3.3.4. Case-by-Case Evaluation of PCR Applied to Infectious Diseases
  • 3.4. Intra-Reginal Cooperation.
  • 4 Case Study: The AMB/ATT Program
  • 4.1. Introduction
  • 4.2. Program Description
  • 4.2.1. Objectives
  • 4.2.2. Format
  • 4.3. Program Structure
  • 4.3.1. Phase
  • 4.3.2. Phase
  • 4.3.3. Phase
  • 4.4. Program Development
  • 4.5. Examples of Projects
  • 4.6. International Courses
  • II Selected Protocols
  • 5.1. PCR Protocols
  • 5.1.1. Overview
  • 5.1.2. Equipment, Materials, Control, and Procedures Common to All PCR Protocols
  • 5.1.3. Dengue Virus
  • 5.1.4. New World Leishmania
  • 5.1.5. Mycobactrium tuberculosis
  • 5.1.6. Plasmodium falciparum and Plasmodium vivax
  • 5.1.7. Vibrio cholerae
  • 5.1.8. Diarrheagenic E. coli and Shigella
  • 5.1.9. Chlamydia trachomatis and Neisseria gonorrhoeae
  • 5.1.10. Leptospira
  • 5.1.11. Trypanosoma cruzi
  • 5.2. Nonradioactive DNA Probes: V. cholerae Colony Blot
  • 5.2.1. Labeling the Probe Using PCR
  • 5.2.2. Preparation of the Colony Blot
  • 5.2.3. Hybridization
  • 5.2.4. Visualization
  • 6 Rapid Cloning of PCR Products
  • 6.1. Primer Design.
  • 6.2. Preparation of PCR Products for Cloning
  • 6.2.1. Digestion of Vector and PCR Products
  • 6.2.2. Preparative Agarose Gel
  • 6.2.3. Purification of the Excised DNA Fragments Using Silica Particles
  • 6.3. Ligation
  • 6.3.1. Checking DNA Fragment Concentration by Agarose Gel Electrophoresis
  • 6.3.2. Ligation Reaction
  • 6.4. Preparation of Competent Cells and Transformation
  • 6.4.1. Preparation of Competent Cells
  • 6.4.2. Transformation
  • 6.5. Checking Clones by PCR
  • 6.6. Plasmid Purification
  • 6.7. Analysis of Clones by Restriction Enzyme Digestion
  • 6.8. Agarose Gel Electrophoresis
  • III Appendix
  • A. Construction of Laboratory Equipment
  • B. In-House Preparation of Reagents
  • B.1 Useful Formulas
  • B.2 Solutions
  • B.3 Preparation of Selected Reagents
  • B.4 DNA Size Markers
  • C. Inventory for a PCR Laboratory
  • D. Good Laboratory Practice
  • D.1 General Tips
  • D.2 Calibration of Adjustable Pipettors
  • E. Prevention of Cross-Contamination
  • F. PCR Troubleshooting Guide and Flow-Chart
  • G. Workshop Organization and Teaching Tips
  • H. Sample Charts and Worksheets
  • I. Useful World Wide Web Sites.

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