Team:KAIT Japan/Notebook

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During the creation.
Creating parts of Tar methylation region.

Contents

Date:8/9

Colony PCR

Reagent

  • TaKaRa Ex Taq(5units/μL) 0.5μL
  • 10×Ex Taq buffer 10μL
  • dNTP Mixture(2.5Meach) 8μL
  • Primer F(10μM) 4μL
  • Primer R(10μM) 4μL
  • Template(E.coli DH5α)

→Reflection:Took colony too many.You should take less colony.
Conditions of the thermal cycler

  1. 95℃(5min)
  2. 94℃(30sec)
  3. 61℃(30sec)
  4. 71℃(40sec)
  5. 72℃(1min)
  6. 4℃(Save)
    • 2~4:35cycle→Reflection:The number of cycles was less.So,We increased The number of cycles in 8/11.(50 cycles)

Date:8/11

The purified DNA

  1. Electrophoresis
    • Marker:pigment(buffer) 1μL,DNA molecule 2μL,TE buffer 3μL
    • Sample:pigment(buffer) 1μL,sample 5μL
    • Gel concentration:1.2%,Migration time:30min

→Reflection:Band was less.

  1. Storage

PCR Production

  1. Electrophoresis
    • The gel check and cut
  2. DNA purification
  3. Confirmation of electrophoresis
  4. PCR
    • 50cycle
  5. Storage

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