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Team:HokkaidoU Japan/Notebook/Pre experiment/mRFP-expression-construct
From 2014.igem.org
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<div class="step-recipe"> R0040-B0034-E1010-B0015 from our stock | <div class="step-recipe"> R0040-B0034-E1010-B0015 from our stock | ||
100up_F Tm:70.4℃ & 200dn_R Tm:66.6℃ | 100up_F Tm:70.4℃ & 200dn_R Tm:66.6℃ | ||
| + | </div> | ||
| + | <div class="step-recipe"> | ||
KOD plus Neo</div> | KOD plus Neo</div> | ||
</div> | </div> | ||
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</ul> | </ul> | ||
<div class="step-recipe"> Cut R0040-B0034-E1010-B0015 with EcoRI, PstI (using 10×H buffer) | <div class="step-recipe"> Cut R0040-B0034-E1010-B0015 with EcoRI, PstI (using 10×H buffer) | ||
| + | </div> | ||
| + | <div class="step-recipe"> | ||
Cut pSB6A1 with EcoRI, PstI (using 10×H buffer) | Cut pSB6A1 with EcoRI, PstI (using 10×H buffer) | ||
</div> | </div> | ||
Revision as of 03:52, 18 October 2014
Notebook
Lab Documents
mRFP Expression Plasmid Construction
-
Start
-
Get genes
pSB6A1 -
Transformation
pSB6A1 1µL to JM109 -
Liquid Culture
pSB6A1 -
Mini-prep
pSB6A1 -
PCR & PCR purification
R0040-B0034-E1010-B0015 from our stock 100up_F Tm:70.4℃ & 200dn_R Tm:66.6℃KOD plus Neo -
Digestion
Cut R0040-B0034-E1010-B0015 with EcoRI, PstI (using 10×H buffer)Cut pSB6A1 with EcoRI, PstI (using 10×H buffer) -
Gel Extraction
R0040-B0034-E1010-B0015 pSB6A1 -
Ethanol precipetation
R0040-B0034-E1010-B0015 pSB6A1 -
Dephosphorylation
R0040-B0034-E1010-B0015 -
Ligation
Ligate R0040-B0034-E1010-B0015 with pSB6A1 -
Transformation
R0040-B0034-E1010-B0015(on pSB6A1) 5µL to DH5α Turbo -
Liquid Culture
R0040-B0034-E1010-B0015(on pSB6A1) -
Mini-prep
R0040-B0034-E1010-B0015(on pSB6A1) -
Complete!
