IGEM:Peking/2007/Count-Conjugation-Notebook/2007-8-2: Difference between revisions
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[[Image:Peking_2007-8-2_F-OriT_digesting_purification_result.jpg]] | [[Image:Peking_2007-8-2_F-OriT_digesting_purification_result.jpg]] | ||
==Ligation:F-OriT ->pSB1A2== | |||
*Ligate the F-Ori-T fragment and pSB1A2 vector | |||
*Ligation system contains (totally 10uL):3uL Ori-T, 2uL pSB1A2, 1uL T4-Ligase, 1uL 10Xbuffer, 3uL dH20 | |||
*4℃ overnight | |||
Revision as of 21:29, 2 August 2007
- OriT by Qu Mingzhi
mini-prep F-OriT_pEasy-T3
- using Transgen mini plasmid puriflication kit.
mini-prep result
- from left to right:
- mini-prep F-OriT_pEasy-T3-1
- mini-prep F-OriT_pEasy-T3-2
- marker
mini-prep double digesting test result
- Digesting F-OriT_pEasy-T3 with EcoR1/PstI.
- Digestion system contains (totally 10uL): 1uL 10*H, 0.25uL EcoRI, 0.25uL PstI, 5uL plasmid, 3.5uL ddH2O.
- 37℃ culutre for 3 hours.
- from left to right:
- mini-prep F-OriT_pEasy-T3-1 @ EcoRI/PstI
- mini-prep F-OriT_pEasy-T3-2 @ EcoRI/PstI
- marker 1
- marker 2
- I think the channel is bright enough for purification,so.... ^_^
Ori-T purification
- use Transgen kit.
- 30uL after purflication
electrophorsis result
- from left to right:
- purified F-Ori-T
- marker
Ligation:F-OriT ->pSB1A2
- Ligate the F-Ori-T fragment and pSB1A2 vector
- Ligation system contains (totally 10uL):3uL Ori-T, 2uL pSB1A2, 1uL T4-Ligase, 1uL 10Xbuffer, 3uL dH20
- 4℃ overnight
