Team:HokkaidoU Japan/Notebook/aggregation Week 11
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+ | <div class="hokkaidou-notebook-daily"> | ||
+ | ==September 10th== | ||
+ | <div> | ||
+ | ==Digestion of eCFP-RBS-pSB1A2 and pBAD-RBS-pSB1A2== | ||
+ | <p> | ||
+ | To make a construct of pBAD-RBS-eCFP-RBS-Ag43-dT-pSTV28 by 3piece ligation, we digested pBAD-RBS-eCFP-RBS-pSB1A2 with EcoRI & SpeI, Ag43-dT-pSB1AK3 with XbaI & NotI and pSTV28 with EcoRI and NotI. Then we digested pT7-RBS-pSB1C3 with XbaI & SpeI as a control for confirmation of the ability of restriction enzyme. | ||
+ | Insert1 (pBAD-RBS-eCFP-RBS-pSB1A2) | ||
+ | {|class="hokkaidou-table-digestion" | ||
+ | |- | ||
+ | |DNA solution ( 30ng/ul) | ||
+ | |22 ul | ||
+ | |- | ||
+ | |XbaI | ||
+ | |1 ul | ||
+ | |- | ||
+ | |PstI | ||
+ | |1 ul | ||
+ | |- | ||
+ | |10xM buffer | ||
+ | |3 ul | ||
+ | |- | ||
+ | |DW | ||
+ | |3 ul | ||
+ | |- | ||
+ | |Total | ||
+ | |30 ul | ||
+ | |} | ||
+ | |||
+ | |||
+ | |||
+ | |||
+ | Vector(pBAD-RBS-pSB1A2) | ||
+ | {|class="hokkaidou-table-digestion" | ||
+ | |- | ||
+ | |DNA solution ( 50ng/ul) | ||
+ | |3 ul | ||
+ | |- | ||
+ | |SpeI | ||
+ | |1 ul | ||
+ | |- | ||
+ | |PstI | ||
+ | |1 ul | ||
+ | |- | ||
+ | |10xH buffer | ||
+ | |2 ul | ||
+ | |- | ||
+ | |DW | ||
+ | |13 ul | ||
+ | |- | ||
+ | |Total | ||
+ | |20 ul | ||
+ | |} | ||
+ | |||
+ | |||
+ | control (eCFP-pSB1A2) | ||
+ | {|class="hokkaidou-table-digestion" | ||
+ | |- | ||
+ | |DNA solution (30 ng/ul) | ||
+ | |5 ul | ||
+ | |- | ||
+ | |XbaI | ||
+ | |1 ul | ||
+ | |- | ||
+ | |SpeI | ||
+ | |1 ul | ||
+ | |- | ||
+ | |10xM buffer | ||
+ | |2 ul | ||
+ | |- | ||
+ | |DW | ||
+ | |11 ul | ||
+ | |- | ||
+ | |Total | ||
+ | |20 ul | ||
+ | |} | ||
+ | |||
+ | |||
+ | |||
+ | {|class="hokkaidou-table-digestion" | ||
+ | |- | ||
+ | |Number | ||
+ | |Degree | ||
+ | |Minute | ||
+ | |- | ||
+ | |1 | ||
+ | |37 | ||
+ | |120 | ||
+ | |- | ||
+ | |2 | ||
+ | |60 | ||
+ | |15 | ||
+ | |- | ||
+ | |3 | ||
+ | |4 | ||
+ | |HOLD | ||
+ | |} | ||
+ | </div></div> | ||
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Revision as of 05:04, 10 September 2012
September 10th
Digestion of eCFP-RBS-pSB1A2 and pBAD-RBS-pSB1A2
To make a construct of pBAD-RBS-eCFP-RBS-Ag43-dT-pSTV28 by 3piece ligation, we digested pBAD-RBS-eCFP-RBS-pSB1A2 with EcoRI & SpeI, Ag43-dT-pSB1AK3 with XbaI & NotI and pSTV28 with EcoRI and NotI. Then we digested pT7-RBS-pSB1C3 with XbaI & SpeI as a control for confirmation of the ability of restriction enzyme. Insert1 (pBAD-RBS-eCFP-RBS-pSB1A2)
DNA solution ( 30ng/ul) | 22 ul |
XbaI | 1 ul |
PstI | 1 ul |
10xM buffer | 3 ul |
DW | 3 ul |
Total | 30 ul |
Vector(pBAD-RBS-pSB1A2)
DNA solution ( 50ng/ul) | 3 ul |
SpeI | 1 ul |
PstI | 1 ul |
10xH buffer | 2 ul |
DW | 13 ul |
Total | 20 ul |
control (eCFP-pSB1A2)
DNA solution (30 ng/ul) | 5 ul |
XbaI | 1 ul |
SpeI | 1 ul |
10xM buffer | 2 ul |
DW | 11 ul |
Total | 20 ul |
Number | Degree | Minute |
1 | 37 | 120 |
2 | 60 | 15 |
3 | 4 | HOLD |
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