Team:Tokyo Metropolitan/Notebook/Fiber/2010/10/24
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+ | __NOTOC__ | ||
+ | ==2010/10/24 Sunday (Naoto)== | ||
+ | :'''member''' | ||
+ | |||
+ | :naoto and watachin | ||
+ | |||
+ | ===Preparation:making preculture for Miniprep === | ||
+ | :'''material''' | ||
+ | |||
+ | :*colony of ''E.coli'' | ||
+ | :**bcsA→No.21,22 | ||
+ | :**bcsB→No.10 | ||
+ | :**bcsC→No.54,64 | ||
+ | :**bcsD→No.75 | ||
+ | :*LB+Cam broth | ||
+ | |||
+ | :'''procedure''' | ||
+ | |||
+ | :see <html><a href="https://2010.igem.org/Team:Tokyo_Metropolitan/Project/Fiber/Protocol#.E3.80.80Grow up culture of E.coli">protocol2</a></html> | ||
+ | |||
+ | ===Experiment:Ligation=== | ||
+ | :'''material''' | ||
+ | |||
+ | :see <html><a href="https://2010.igem.org/Team:Tokyo_Metropolitan/Project/Fiber/Protocol#.E3.80.80ligation">protocol8</a></html> | ||
+ | :*vector | ||
+ | :**pUC19(digested by XbaI) | ||
+ | :*Insert | ||
+ | :**bcsA(digested by XbaI+SpeI) | ||
+ | :**bcsB(digested by XbaI+SpeI) | ||
+ | :**bcsC(digested by XbaI+SpeI) | ||
+ | :**bcsD(digested by XbaI+SpeI) | ||
+ | |||
+ | :'''procedure''' | ||
+ | |||
+ | :refer to <html><a href="https://2010.igem.org/Team:Tokyo_Metropolitan/Project/Fiber/Protocol#.E3.80.80ligation">protocol8</a></html> | ||
+ | |||
+ | ===Experiment:Transformation=== | ||
+ | |||
+ | :'''material''' | ||
+ | |||
+ | :see <html><a href="https://2010.igem.org/Team:Tokyo_Metropolitan/Project/Fiber/Protocol#.E3.80.80transformation">protocol9</a></html> | ||
+ | :*Ecos JM109/Nova Blue(Competent Cell) | ||
+ | |||
+ | :'''procedure''' | ||
+ | |||
+ | :refer to <html><a href="https://2010.igem.org/Team:Tokyo_Metropolitan/Project/Fiber/Protocol#.E3.80.80transformation">protocol9</a></html> | ||
+ | |||
+ | ===Experiment:Miniprep=== | ||
+ | :'''material''' | ||
+ | |||
+ | :see <html><a href="https://2010.igem.org/Team:Tokyo_Metropolitan/Project/Fiber/Protocol#.E3.80.80miniprep">protocol10</a></html> | ||
+ | :*Preculture | ||
+ | :No.21,22:bcsA | ||
+ | :No.10:bcsB | ||
+ | :No.54,64:bcsC | ||
+ | :No.75:bcsD | ||
+ | |||
+ | :'''procedure''' | ||
+ | |||
+ | :refer to <html><a href="https://2010.igem.org/Team:Tokyo_Metropolitan/Project/Fiber/Protocol#.E3.80.80miniprep">protocol10</a></html> | ||
+ | |||
+ | ===Experiment:Sequence=== | ||
+ | :'''material''' | ||
+ | :*PCR | ||
+ | :**DNA(No.21,22,10,54,64,75:following before "Miniprep") | ||
+ | :**Big Dye | ||
+ | :**primer | ||
+ | :**DW | ||
+ | :*Ethanol precipitation | ||
+ | :**ethanol | ||
+ | :**EDTA | ||
+ | |||
+ | :'''procedure''' | ||
+ | :#mix materials(DNA<50ng) | ||
+ | :#PCR | ||
+ | :#Add EDTA and Ethanol and put at room temperature (15min) | ||
+ | :#centrifuge (8000rpm,30min) | ||
+ | :#throw away supernatant | ||
+ | :#add ethanol again | ||
+ | :#centrifuge (8000rpm,15min) | ||
+ | :#put at refrigerator | ||
+ | :#throw away supernatant | ||
+ | :#add Hi-Di solution | ||
+ | :#transfer these sample to plate for sequence | ||
+ | :#read sequence | ||
+ | |||
+ | |||
+ | |||
+ | <br/> |
Latest revision as of 18:08, 27 October 2010
E.coli Fiber Project Notebook
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|
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2010/10/24 Sunday (Naoto)
- member
- naoto and watachin
Preparation:making preculture for Miniprep
- material
- colony of E.coli
- bcsA→No.21,22
- bcsB→No.10
- bcsC→No.54,64
- bcsD→No.75
- LB+Cam broth
- colony of E.coli
- procedure
- see protocol2
Experiment:Ligation
- material
- see protocol8
- vector
- pUC19(digested by XbaI)
- Insert
- bcsA(digested by XbaI+SpeI)
- bcsB(digested by XbaI+SpeI)
- bcsC(digested by XbaI+SpeI)
- bcsD(digested by XbaI+SpeI)
- vector
- procedure
- refer to protocol8
Experiment:Transformation
- material
- see protocol9
- Ecos JM109/Nova Blue(Competent Cell)
- procedure
- refer to protocol9
Experiment:Miniprep
- material
- see protocol10
- Preculture
- No.21,22:bcsA
- No.10:bcsB
- No.54,64:bcsC
- No.75:bcsD
- procedure
- refer to protocol10
Experiment:Sequence
- material
- PCR
- DNA(No.21,22,10,54,64,75:following before "Miniprep")
- Big Dye
- primer
- DW
- Ethanol precipitation
- ethanol
- EDTA
- PCR
- procedure
- mix materials(DNA<50ng)
- PCR
- Add EDTA and Ethanol and put at room temperature (15min)
- centrifuge (8000rpm,30min)
- throw away supernatant
- add ethanol again
- centrifuge (8000rpm,15min)
- put at refrigerator
- throw away supernatant
- add Hi-Di solution
- transfer these sample to plate for sequence
- read sequence