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Team:HokkaidoU Japan/Notebook/asB0034/asE1010 1A3 Construction
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<li class="ldd_heading"><a href="https://2014.igem.org/Team:HokkaidoU_Japan/Outreach/Discussion">Discussion</a></li> | <li class="ldd_heading"><a href="https://2014.igem.org/Team:HokkaidoU_Japan/Outreach/Discussion">Discussion</a></li> | ||
<li class="ldd_contents"><a href="https://2014.igem.org/Team:HokkaidoU_Japan/Outreach/Discussion#Background">Background</a></li> | <li class="ldd_contents"><a href="https://2014.igem.org/Team:HokkaidoU_Japan/Outreach/Discussion#Background">Background</a></li> | ||
| - | <li class="ldd_contents"><a href="https://2014.igem.org/Team:HokkaidoU_Japan/Outreach/Discussion# | + | <li class="ldd_contents"><a href="https://2014.igem.org/Team:HokkaidoU_Japan/Outreach/Discussion#Evaluation">Evaluation</a></li> |
</ul> | </ul> | ||
</div> | </div> | ||
Revision as of 02:49, 18 October 2014
Notebook
Lab Documents
Anti-Sense E1010 on 1A3 Construction
-
Start
-
PCR
R0010-stem-asE1010-stem-B0015(on pSB1C3) EcoRI_XbaI_F Tm:74.5℃ & Ptet_SpeI_R(insert) Tm:72.6℃ KOD Fx Neo -
Digestion
Cut vector with EcoRI, PstI (using 10×H buffer) Cut insert with EcoRI, PstI (using 10×M buffer) -
Dephosphorylation
Antactic phosphorylate pSB1A3 (using 10×Antarctic phosphatase buffer) 37°C 35min 65℃ 10min -
Ligation
Ligate pSB1A3 with R0010-stem-asE1010-stem-B0015 -
Transformation
R0010-stem-asE1010-stem-B0015(on pSB1A3) 5.0µL to DH5α -
Colony PCR
R0010-stem-asE1010-stem-B0015(on pSB1A3) EcoI_XbaI_F Tm:74.5℃ & PstI_SpeI_R Tm:74.6℃ Kapa-Taq -
Culture
R0010-stem-asE1010-stem-B0015(on pSB1A3) -
Mini-Prep
R0010-stem-asE1010-stem-B0015(on pSB1A3) -
Complete!
