GENETECH 

Course 4: Polymerase Chain Reaction (PCR) Technology (14 hours)

 

This course is suitable for school-level, undergraduate and graduate students, and those engaged in scientific research, diagnostics and teaching which involves Polymerase Chain Reaction (PCR) Technology. 

 

 

Theory:

 

• Guidelines for PCR

• Principle of PCR

• Critical factors for successful PCR

• Designing PCR primers

• Standard practices in a PCR Laboratory

• Troubleshooting and prevention of carryover contamination

• Optimization strategies for PCR

 

     PCR product analysis:

 

• Detection of PCR products by gel electrophoresis

• Methods of purification and quantification of PCR products

• Methods of labelling of PCR products (PCR probes) and visualization of label

• Cloning of PCR products: TOPO cloning

• DNA sequence analysis of PCR product 

 

     PCR Applications:

 

• Restriction Fragment Length Polymorphism (RFLP) and Randomly Amplified Polymorphic DNA (RAPD) analysis

• Short Tandem Repeat (STR) (micro-satellite) analysis

• Multiplex PCR

• Reverse Transcription (RT)-PCR

• PCR-ELISA (Enzyme Linked Immunosorbent Assay)

• In-situ PCR

• Quality Control/Quality Assurance of PCR assays

 

 

Practicals:

 

• Extraction of DNA from cells, blood stains for PCR

• A standard PCR assay

• Analysis of PCR products by agarose and acrylamide gel electrophoresis

• Strategies for optimization of PCR

• Purification of PCR products

• Labelling of PCR products

• Primer design using online sequence databases

• PCR-RFLP assay

• Multiplex PCR assay

• RT-PCR assay

• PCR-ELISA assay

• PCR troubleshooting

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