IGEM:Peking/2007/Count-Conjugation-Notebook/2007-7-21

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< IGEM:Peking | 2007 | Count-Conjugation-Notebook(Difference between revisions)
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Current revision (10:03, 4 September 2007) (view source)
 
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==transformation e0040->pSB3k3==
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==Ligation e0040->pSB3k3==
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*use T4 Ligase to transform.
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*use T4 Ligase to ligate.
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*Digestion system contains (totally 20uL): 4uL pSB3K3(I7100), 3uLEGFP(E0040),1uL 10XT4L buffer, 1uL T4 L, 1uL dH20.
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*Ligation system contains (totally 20uL): 4uL pSB3K3(I7100), 3uLEGFP(E0040),1uL 10XT4L buffer, 1uL T4 L, 1uL dH20.
===PCR crRNA===
===PCR crRNA===

Current revision

Ligation e0040->pSB3k3

  • use T4 Ligase to ligate.
  • Ligation system contains (totally 20uL): 4uL pSB3K3(I7100), 3uLEGFP(E0040),1uL 10XT4L buffer, 1uL T4 L, 1uL dH20.

PCR crRNA

  • PCR system contains (totally 10uL): 0.5uL P1, 0.5uL P2, 2.5uL 10*b, dNTP 2uL, Taq 0.25uL, dH20 19.25uL.
  • PCR condition:
  1. 94℃ 5min
  2. 94℃ 30s
  3. 60℃ 30s
  4. 72℃ 30s
  5. Go to step 2 for 29 times
  6. 72℃ 10min
  7. 4℃ end
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