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Team:HokkaidoU Japan/Notebook/asB0034/asB0034-construction
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Revision as of 23:51, 17 October 2014
Notebook
Lab Documents
asB0034 Construction
-
Start
-
Primer Annealing
asB0034_F Tm: 68.7℃ asB0034_R Tm: 68.7℃ -
Digestion
Cut pHN1257 with NcoI, XhoI(using 10×cut smart) -
Gel extraction
digested pHN1257 -
Ethanol Precipetation
digested pHN1257 -
Ligation
Ligate pHN1257 with asB0034 -
Transformation
Ptrc-stem-asB0034-stem(pHN1257) on ice 30min each of 5.0µL DNA to DH5α 42℃ 1min on ice 2min -
Colony PCR
Ptrc-stem-asB0034-stem(pHN1257) pHN1257_up_F Tm:60.1℃ asB0034_R Tm:68.7℃ -
Culture
Ptrc-stem-asB0034-stem -
mini-Prep
Ptrc-stem-asB0034-stem -
Transformation
Ptrc-stem-asB0034-stem on pHN1257&Ptet-B0034-mRFP-dt on pSB6A1 on ice 30min each of 2.5µL DNA to DH5α 42℃ 30sec on ice 2min -
Complete!
