Molecular Diagnostics

POLYMERASE CHAIN REACTION 

• DENATURATION – 94 – 95 degree Celsius (15 – 30 secs) 
  
  • For dsDNA separation 
• PRIMER ANNEALING – 45 – 65 degree Celsius (30 secs. – 2 mins.) 
  
  • Anneals primer to target DNA
• PRIMER EXTENSION – 68 – 72 degree Celsius 
  • Synthesis of new strands of DNA

PCR COMPONENTS 

• Template DNA – target for PCR 
• Oligonucleotide Primers – starts synthesis new strands of DNA 
• Thermostable DNA – synthesizes new strands of polymerase of DNA 
• Magnesium – required by DNA polymerase for proper reaction 
• Buffer – ensures proper conditions and pH for DNA polymerase 
• Deoxynucleotides – used by polymerase to synthesize new DNA 
• Thermal Cycler – heats and cools PCR cycle steps