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Team:Evry/Notebook/Sensing/PCBs/08-21-2014
From 2014.igem.org
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| - | <u>Sensor construction | + | <u>Sensor construction bphR2/PbpR1</u> |
<br/><a href="parts.igem.org/Part:BBa_J23114">BBa_J23114</a> was resuspended with 10 µL sterile water in order to obtain a DNA concentration around 0.2 ng/µl (according to the registry). Solution was transferred into one 1 ml eppendorf tube and stored at -20°C. | <br/><a href="parts.igem.org/Part:BBa_J23114">BBa_J23114</a> was resuspended with 10 µL sterile water in order to obtain a DNA concentration around 0.2 ng/µl (according to the registry). Solution was transferred into one 1 ml eppendorf tube and stored at -20°C. | ||
<br/>Transformations of constitutive promoter <a href="parts.igem.org/Part:BBa_J23114">BBa_J23114</a>, RFP <a href="parts.igem.org/Part:BBa_E1010">BBa_E1010</a> and terminator <a href="parts.igem.org/Part:BBa_B0015">BBa_B0015</a> and vector pSB1C3G3 was done on DH5alpha as followed: </p> | <br/>Transformations of constitutive promoter <a href="parts.igem.org/Part:BBa_J23114">BBa_J23114</a>, RFP <a href="parts.igem.org/Part:BBa_E1010">BBa_E1010</a> and terminator <a href="parts.igem.org/Part:BBa_B0015">BBa_B0015</a> and vector pSB1C3G3 was done on DH5alpha as followed: </p> | ||
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<br/>pSB1C3 was digested with EcoRI and pstI and a ligation with bphR2 was done before the transformation in DH5a. | <br/>pSB1C3 was digested with EcoRI and pstI and a ligation with bphR2 was done before the transformation in DH5a. | ||
<br/> 50 µL were plated on Cam Lb plate and incubated at 37°C overnight | <br/> 50 µL were plated on Cam Lb plate and incubated at 37°C overnight | ||
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<span class="cd-date">Aug 21</span> | <span class="cd-date">Aug 21</span> | ||
Latest revision as of 00:15, 13 October 2014
Sensor construction bphR2/PbpR1
BBa_J23114 was resuspended with 10 µL sterile water in order to obtain a DNA concentration around 0.2 ng/µl (according to the registry). Solution was transferred into one 1 ml eppendorf tube and stored at -20°C.
Transformations of constitutive promoter BBa_J23114, RFP BBa_E1010 and terminator BBa_B0015 and vector pSB1C3G3 was done on DH5alpha as followed:
- Remove E. coli competent tubes from -80°C and keep it on ice
- Add 3 µl of template (here solubilized plasmids from the registry distribution kit) and mix gently
- Incubate 10 minutes on ice
- Perform an heat shock 30 seconds at 42°C
- Incubate 2 minutes on ice
- Add 2 ml of LB medium and incubate 60 minutes at 37°C with an agitation at 200 rpm
- Plate 200 µl of pSB1C3G3, BBa_B0015 and E1010 on a chloramphenicol LB agar plate and ampicilline LB agar plate for BBa_J23114
- Incubate plate overnight at 37°C
We received primers.
pSB1C3 was digested with EcoRI and pstI and a ligation with bphR2 was done before the transformation in DH5a.
50 µL were plated on Cam Lb plate and incubated at 37°C overnight
Aug 21
