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

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