Team:HokkaidoU Japan/Notebook/Promoter WeakPromoter-I732020
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Latest revision as of 16:41, 27 September 2013
Maestro E.coli
Notebook
Weak Promoter-I732020
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Start
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Transformation
J23112, J23103, J23117, J23106, J23111 from distribution kit
each of 1µL DNA to DH5α -
Culture
J23112, J23103, J23117, J23106, J23111
I732020 from our stock -
Mini-prep
J23112, J23103, J23117, J23106, J23111, I732020 -
PCR
I732020
100up, 200dn, KOD Plus Neo -
Digestion
Cut (J23112, J23103, J23117, J23106, J23111) with SpeI, PstI (using 10×H buffer)
Cut I732020 with XbaI, PstI (using 10×M buffer) -
Gel extraction
J23112, J23103, J23117, J23106, J23111 -
Antarctic phosphatase
J23112, J23103, J23117, J23106, J23111 -
Ligation
Ligate (J23112, J23103, J23117, J23106, J23111) with I732020 -
Transformation
(J23112, J23103, J23117, J23106, J23111)-I732020
each of 1µL DNA to DH5α -
Colony PCR & Culture
(J23112, J23103, J23117, J23106, J23111)-I732020 -
Mini-prep
(J23112, J23103, J23117, J23106, J23111)-I732020 -
PCR
(J23112, J23103, J23117, J23106, J23111)-I732020
EX-f, PS-r, KOD FX NEO -
PCR purification
(J23112, J23103, J23117, J23106, J23111)-I732020 -
Digestion
Cut K1084007 on pSB1C3 with EcoRI, PstI (using 10×H buffer)
Cut (J23112, J23103, J23117, J23106, J23111)-I732020 with EcoRI, PstI (using 10×H buffer) -
Antarctic phophatase
pSB1C3 -
Ligation
Ligate pSB1C3 with (J23112, J23103, J23117, J23106, J23111)-I732020 -
Transformation
(J23112, J23103, J23117, J23106, J23111)-I732020 -
Complete!