"
Team:HokkaidoU Japan/Notebook/plastic Week 13
From 2012.igem.org
(Difference between revisions)
| (2 intermediate revisions not shown) | |||
| Line 7: | Line 7: | ||
<div class="hokkaidou-section"> | <div class="hokkaidou-section"> | ||
| - | ====Single | + | ====Single colony isolation==== |
We got few colonies from the plate. | We got few colonies from the plate. | ||
The colonies were isolated to another plate. | The colonies were isolated to another plate. | ||
| Line 13: | Line 13: | ||
<div class="hokkaidou-notebook-daily"> | <div class="hokkaidou-notebook-daily"> | ||
| + | |||
===September 25th=== | ===September 25th=== | ||
<div class="hokkaidou-section"> | <div class="hokkaidou-section"> | ||
| Line 20: | Line 21: | ||
====PCR==== | ====PCR==== | ||
| - | |||
Did PCR for promoter of R.eutropha from pGEM to ligate with RBS-PhaC-RBS-PhaA-RBS-PhaB-dT on pSB1A2. | Did PCR for promoter of R.eutropha from pGEM to ligate with RBS-PhaC-RBS-PhaA-RBS-PhaB-dT on pSB1A2. | ||
| - | |||
====Digestion==== | ====Digestion==== | ||
| - | |||
Digested the promoter done PCR and RBS-PhaA on pSB1A2 with XbaI and SpeI, and plasmid of RBS-PhaC-RBS-PhaA-RBS-PhaB-dT on pSB1A2 with XbaI. | Digested the promoter done PCR and RBS-PhaA on pSB1A2 with XbaI and SpeI, and plasmid of RBS-PhaC-RBS-PhaA-RBS-PhaB-dT on pSB1A2 with XbaI. | ||
| - | |||
====Gel extraction==== | ====Gel extraction==== | ||
| - | |||
We confirmed succession of digestion by electrophoresis. | We confirmed succession of digestion by electrophoresis. | ||
And then DNA were extracted from TBE gel. | And then DNA were extracted from TBE gel. | ||
| - | + | ||
| + | ====Ethanol precipitation==== | ||
| + | The digested DNAs were condensed by Ethanol precipitation. | ||
====Ligation==== | ====Ligation==== | ||
| - | |||
Ligated the promoter with RBS-PhaC-RBS-PhaA-RBS-PhaB-dT and with pSB1A2. | Ligated the promoter with RBS-PhaC-RBS-PhaA-RBS-PhaB-dT and with pSB1A2. | ||
| - | |||
====Transformation==== | ====Transformation==== | ||
| - | |||
Transformed each ligation product into JM109, and incubated. | Transformed each ligation product into JM109, and incubated. | ||
| - | |||
</div></div> | </div></div> | ||
<div class="hokkaidou-notebook-daily"> | <div class="hokkaidou-notebook-daily"> | ||
| + | |||
===September 26th=== | ===September 26th=== | ||
<div class="hokkaidou-section"> | <div class="hokkaidou-section"> | ||
====Colony PCR==== | ====Colony PCR==== | ||
| - | |||
Colony PCR for colonies transformed at 25th. | Colony PCR for colonies transformed at 25th. | ||
| - | |||
</div></div> | </div></div> | ||
| + | |||
<!-- DO NOT EDIT UNDER THIS LINE @iTakeshi --> | <!-- DO NOT EDIT UNDER THIS LINE @iTakeshi --> | ||
</div> | </div> | ||
<br style="line-height: 0; clear: both;" /> | <br style="line-height: 0; clear: both;" /> | ||
{{Team:HokkaidoU_Japan/footer}} | {{Team:HokkaidoU_Japan/footer}} | ||
Latest revision as of 22:27, 26 September 2012
Contents |
September 24th
Single colony isolation
We got few colonies from the plate. The colonies were isolated to another plate.
September 25th
Liquid culture
We got colonies on the plate, and started liquid culture.
PCR
Did PCR for promoter of R.eutropha from pGEM to ligate with RBS-PhaC-RBS-PhaA-RBS-PhaB-dT on pSB1A2.
Digestion
Digested the promoter done PCR and RBS-PhaA on pSB1A2 with XbaI and SpeI, and plasmid of RBS-PhaC-RBS-PhaA-RBS-PhaB-dT on pSB1A2 with XbaI.
Gel extraction
We confirmed succession of digestion by electrophoresis. And then DNA were extracted from TBE gel.
Ethanol precipitation
The digested DNAs were condensed by Ethanol precipitation.
Ligation
Ligated the promoter with RBS-PhaC-RBS-PhaA-RBS-PhaB-dT and with pSB1A2.
Transformation
Transformed each ligation product into JM109, and incubated.
September 26th
Colony PCR
Colony PCR for colonies transformed at 25th.
