IGEM:Peking/2007/Count-Conjugation-Notebook/2007-7-31: Difference between revisions
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==Tandem OriT== | ==Tandem OriT== | ||
*by Qu Mingzhi | *by Qu Mingzhi | ||
*get primer | *get F-OriT primer, primer vf,vf2,vr,pf1,pr1 stored as ~100uM. | ||
*PCR system contains (totally 50uL):1uL Primer pf1, 1uL Primer pr1, 4uL dNTP, 0.5uL Taq, 1uL OriT template, 37.5uL dH20, 5uL 10X buffer, | |||
*use different PCR program to test efficiency. | |||
#PCR program condition 1: 94℃ 5min, 94℃ 30s, 51℃ 30s, 72℃ 30s, Go to step 2 for 29 times, | |||
72℃ 10min, 4℃ end. | |||
#PCR program condition 2: 94℃ 5min, 94℃ 30s, 55℃ 30s, 72℃ 30s, Go to step 2 for 29 times, | |||
72℃ 10min, 4℃ end . | |||
*Primer final concentration 1uM. | |||
===electrophorsis result=== | |||
*from ''left'' to ''right'': | |||
==PCR OriT== |
Revision as of 05:54, 1 August 2007
Tandem OriT
- by Qu Mingzhi
- get F-OriT primer, primer vf,vf2,vr,pf1,pr1 stored as ~100uM.
- PCR system contains (totally 50uL):1uL Primer pf1, 1uL Primer pr1, 4uL dNTP, 0.5uL Taq, 1uL OriT template, 37.5uL dH20, 5uL 10X buffer,
- use different PCR program to test efficiency.
- PCR program condition 1: 94℃ 5min, 94℃ 30s, 51℃ 30s, 72℃ 30s, Go to step 2 for 29 times,
72℃ 10min, 4℃ end.
- PCR program condition 2: 94℃ 5min, 94℃ 30s, 55℃ 30s, 72℃ 30s, Go to step 2 for 29 times,
72℃ 10min, 4℃ end .
- Primer final concentration 1uM.
electrophorsis result
- from left to right: