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Polymerase Chain Reaction (PCR) to Amplify rRNA Gene Fragment

1. Prepare sufficient master mix for both partners (45 mL/50 mL reaction)
• 10 mL 10x PCR buffer
• 10 mL 2.5 mM dNTPs (0.25 mM final concentration)
• 15 mL Primer A (5 pmole/mL)
• 15 mL Primer B (5 pmole/mL)
• 40 mL H₂O
• 0.4 mL TaKaRa Ex-Taq DNA Polymerase (2 units)(Panvera)
• 90 mL
  
  
2. Aliquot 45 mL of master mix into each of two 0.5 mL microfuge tubes labelled with the student's initials; PCR; date.
   
   
3. Add 1-5 mL template DNA¹ (100 ng for bacterial genomic DNA), 0-4 mL H₂O to yield a final volume of 50 mL.
   
   
4. Ad