Team:Exeter/lab book/gibs/wk5

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         <a href="https://2012.igem.org/Team:Exeter/lab_book/gibs/wk10"; style="color:#1d1d1b">10th - 14th September</a>
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         <a href="https://2012.igem.org/Team:Exeter/Results#usepoly"; style="color:#e30614" target="_blank"><font size="3"><b>Results: Part 1</b></font></a>
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         <a href="https://2012.igem.org/Team:Exeter/lab_book/gibs/wk11"; style="color:#1d1d1b">17th - 21st September</a>
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         <a href="https://2012.igem.org/Team:Exeter/Results/GvsB"; style="color:#e30614" target="_blank"><font size="3"><b>Results: Part 2</b></font></a>
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    <p><u>Website Designed and Built by: Ryan Edginton, James Lynch & Alex Clowsley</u> &nbsp;&nbsp;|&nbsp;&nbsp;
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    <a href="https://igem.org/Team.cgi?id=764" style="color:#57B947" target="_blank"><u>Contact Us</u></a>  &nbsp;&nbsp;|&nbsp;&nbsp;
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    <a href="https://2012.igem.org/Team:Exeter/site_map" style="color:#57B947"><u>Site Map</u></a></p>
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Latest revision as of 00:10, 27 September 2012

ExiGEM2012 Lab Book Gibs wk5

Operon Construction: 6th - 10th August 2012

 **Monday 6.8.12**


3A BioBrick assembly of RBS-wbbC(d) (BBa_B0034 + BBa_K764005)


Upstream

RBS 3 (BBa_B0034) plasmid DNA - 8.1 µl

Water - 14.4 µl


Downstream wbbC(d)

DNA - 2.8 µl

Water - 19.7 µl


Destination plasmid pSB1C3 (chloramphenicol resistant) Digestion

Ligation


Upstream: 2 µl

Downstream: 2 µl

MilliQ water 2.5


23 °C (room temp.) incubation for 1 hour

20 minute heat kill 80 °C


**Wednesday 8.8.12**


Transformed RBS-wbbC(d) (BBa_B0034 + BBa_K764005)

2 µl into 25 µl competent cells

50 µl and 150 µl LB(chlor) spread plates made


**Thursday 9.8.12**


Transferred RBS-wbbC(d) (BBa_B0034 + BBa_K764005) transformed colonies to LB(chlor) medium


**Friday 10.8.12**


RBS-wbbC(d) transformed cells broth tubes centrifuged for 5 minutes, 3901 rcf, 4 °C

Supernatant discarded

Miniprep according to Fermentas protocol

Step 1045µl milli-Q water added in place of elution buffer, followed by another 5 µl after incubation and centrifugation

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