IGEM:Peking/2007/Count-Conjugation-Notebook/2007-7-18: Difference between revisions
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*EcoR I single digestion. | *EcoR I single digestion. | ||
*Digestion system contains (totally 20uL): 2uL 10*H, 0.5uL EcoR I, 3uL plasmid, 15uL ddH2O. | *Digestion system contains (totally 20uL): 2uL 10*H, 0.5uL EcoR I, 3uL plasmid, 15uL ddH2O. | ||
===electrophoresis result=== | |||
*from ''Left'' to ''Right'': | *from ''Left'' to ''Right'': | ||
#I7100 @ EcoRI | #I7100 @ EcoRI | ||
Line 14: | Line 15: | ||
[[Image:2007-7-18_I7100_E0040_EcoRI.jpg ]] | [[Image:2007-7-18_I7100_E0040_EcoRI.jpg ]] | ||
==Double Digesting E0040 and I7100 for ligation | ==Double Digesting E0040 and I7100 for ligation == | ||
*Method choice test :Oven or Water bath | |||
*Digesting E0040 and I7100 with EcoR1/SpeI and EcoRI/XbaI. | *Digesting E0040 and I7100 with EcoR1/SpeI and EcoRI/XbaI. | ||
*Digestion system contains (totally 20uL): | *Digestion system contains (totally 20uL): | ||
Line 20: | Line 22: | ||
#3uL 10*M, 1uL EcoRI, 1uL XbaI, 6uL plasmid, 10uL ddH2O. | #3uL 10*M, 1uL EcoRI, 1uL XbaI, 6uL plasmid, 10uL ddH2O. | ||
*37℃ culutre for 12 hours. | *37℃ culutre for 12 hours. | ||
===electrophoresis result=== | |||
*from ''Left'' to ''Right'': | *from ''Left'' to ''Right'': | ||
#oven:I7100 @ EcoRI/XbaI | #oven:I7100 @ EcoRI/XbaI | ||
Line 27: | Line 30: | ||
#Marker | #Marker | ||
[[Image:I7100-EcoRI+XbaI_E0040-EcoRI+SpeI.jpg]] | [[Image:I7100-EcoRI+XbaI_E0040-EcoRI+SpeI.jpg]] | ||
===conclusion=== | |||
:Water bath is better. | |||
=OriT Knock Out= | |||
*By Xu Anting and Liu Ting | |||
==Chromosome Extraction== | |||
*Received chromosome of F/R751/pSC101 strain and diluted in 1:50 ddH2O (pH 8.0). | |||
==Travel to Grassland!== | |||
*Everyone at Room 307 in Biotech Building planned to have a travel for three days. All experiment suspended ^o^ |
Latest revision as of 08:44, 21 August 2007
Miniprep
- Miniprep & preservation: I7100.
Digestion Test: I7100 and E0040
- I7100 and E0040.
- EcoR I single digestion.
- Digestion system contains (totally 20uL): 2uL 10*H, 0.5uL EcoR I, 3uL plasmid, 15uL ddH2O.
electrophoresis result
- from Left to Right:
- I7100 @ EcoRI
- E0040 @ EcoRI
- I7100 plasmid
- E0040 plasmid
- Marker
Double Digesting E0040 and I7100 for ligation
- Method choice test :Oven or Water bath
- Digesting E0040 and I7100 with EcoR1/SpeI and EcoRI/XbaI.
- Digestion system contains (totally 20uL):
- 3uL 10*H, 1uL EcoRI, 1uL SpeI, 6uL plasmid, 10uL ddH2O.
- 3uL 10*M, 1uL EcoRI, 1uL XbaI, 6uL plasmid, 10uL ddH2O.
- 37℃ culutre for 12 hours.
electrophoresis result
- from Left to Right:
- oven:I7100 @ EcoRI/XbaI
- Oven:E0040 @ EcoRI/SpeI
- Water Bath: I7100 @ EcoRI/XbaI
- Water Bath: E0040 @ EcoRI/SpeI
- Marker
conclusion
- Water bath is better.
OriT Knock Out
- By Xu Anting and Liu Ting
Chromosome Extraction
- Received chromosome of F/R751/pSC101 strain and diluted in 1:50 ddH2O (pH 8.0).
Travel to Grassland!
- Everyone at Room 307 in Biotech Building planned to have a travel for three days. All experiment suspended ^o^