PCR(Ex-Taq)

Preparation

• 5′primer
• 3′primer
• 2.5mMdNTP
• 10xstanderd　buffer
• MilliQ
• template DNA
• Ex-Taq

Procedure

!use PCR tube

• 1. add these things
  • 0.2μl　100?M5′primer
  • 0.2μl　100?M3′primer
  • 1.6μl　2.5mMdNTP
  • 2μl　10xPfu Ultra 2 buffer
  • ？μl　MilliQ（up to 20μl）
  • ？μl　template DNA（20μl for under 200ng）
• 2. mix by using vortex
• 3. add 0.2μl　Ex-Taq　and dissolve it by flipping
• 4. PCR program
  • 95 degrees / 2min
  • 95 degrees / 30sec
  • 52 to 55 degrees / 30sec （depends on the melting temperature of primer）
  • 72.5 degrees / 1min×（　）kb （the length of DNA you want to increase） ＋ sec as you satisfied 　　　yourself
  • 95 degrees / 30sec (repeat 29 times from here)
  • 25 degrees / ∞