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Imperial College/26 August 2008
From 2008.igem.org
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*A 20ml ''B.subtilis'' culture was prepared in the afternoon. This culture will be used for the microscope tomorrow. | *A 20ml ''B.subtilis'' culture was prepared in the afternoon. This culture will be used for the microscope tomorrow. | ||
*In addition two ''E.coli'' 100ml cultures were grown overnight for midi preping. These ''E.coli'' contained the integration plasmids pDR110 and pDR111. We require these midi preps to PCR out biobricks and for use in transformation protocol optimisation. | *In addition two ''E.coli'' 100ml cultures were grown overnight for midi preping. These ''E.coli'' contained the integration plasmids pDR110 and pDR111. We require these midi preps to PCR out biobricks and for use in transformation protocol optimisation. | ||
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| + | ===Cloning=== | ||
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| + | *Trial PCRs runs with a Pfu enzyme without endonuclease | ||
| + | [[Image:26-8.PNG|center|frame| Lanes : M = Marker, 1 = Vector test (-2°C), 2 = Vector test (-4°C), 3 = Vector test (-6°C), 4 = 100ng Genomic test (-2°C), 5 = 100ng Genomic test (-4°C), 6 = 100ng Genomic test (-6°C), 7 = 1μl Genomic test (-2°C), 8 = 1μl Genomic test (-4°C), 9 = 1μl Genomic test (-6°C), 10 = control (No template DNA)]] | ||
| + | *40ng of vector was used for vector PCRs. Timings were the same as those for our [http://www.openwetware.org/wiki/IGEM:IMPERIAL/2008/New/Protocols/PCR Pfu protocol] | ||
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| + | *For the vector PCRs and the control, primers for the 5' AmyE integration sequence were used. For genomic PCRs, the XylR primers were used | ||
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| + | *Only the vector PCRs were succesful and a contaminent was produced. Further trials will be required | ||
Revision as of 23:04, 1 September 2008
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26 August 2008Wet LabB.subtilis
Cloning
 Lanes : M = Marker, 1 = Vector test (-2°C), 2 = Vector test (-4°C), 3 = Vector test (-6°C), 4 = 100ng Genomic test (-2°C), 5 = 100ng Genomic test (-4°C), 6 = 100ng Genomic test (-6°C), 7 = 1μl Genomic test (-2°C), 8 = 1μl Genomic test (-4°C), 9 = 1μl Genomic test (-6°C), 10 = control (No template DNA)
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