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Team:HokkaidoU Japan/Notebook/Pre experiment/asmRFP
From 2014.igem.org
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Revision as of 02:05, 18 October 2014
Notebook
Lab Documents
Anti-Sense mRFP Construction
-
Start
-
Get anti-sense vector (pHN1257)
Great thanks to N. Nakashima -
PCR & PCR purification
R0040-B0034-E1010-B0015 with as_RBS_XhoI_F Tm:68.2℃ & as_mRFP_NcoI_R Tm:76.0℃ KOD plus Neo -
Ethanol Precipitation
asmRFP -
Digestion
Cut pHN1257 with NcoI, XhoI (using 10×Cut Smart) Cut asmRFP with NcoI, XhoI (using 10×Cut Smart) -
Gel Extraction
pHN1257 asmRFP -
Ethanol Precipitation
asmRFP -
Dephosphorylation
asmRFP -
Ligation
Ligate asmRFP with pHN1257 -
Transformation
asmRFP(on pHN1257) 5µL to DH5α Turbo -
Colony PCR
asmRFP(on pHN1257) pHN1257_up_F Tm:60.1℃ & pHN1257_terminator_R Tm:71.4℃ Kapa-Taq -
Liquid Culture
asmRFP(on pHN1257) -
Mini-Prep
asmRFP(pHN1257) -
Complete!
