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Team:Heidelberg/Templates/MM week19p
From 2013.igem.org
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Nils.kurzawa (Talk | contribs) (Created page with " == 2013-09-02 == * send pIK1.11, pIK1.13, pIK1.14 to sequencing with primer VF2 == 2013-09-03 == * sequences arrived: [[:File:PIK1.11-pIK1.13-pIK1.14-2013-09-03.zip|PIK1.11-pIK...") |
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== 2013-09-02 == | == 2013-09-02 == | ||
* send pIK1.11, pIK1.13, pIK1.14 to sequencing with primer VF2 | * send pIK1.11, pIK1.13, pIK1.14 to sequencing with primer VF2 | ||
== 2013-09-03 == | == 2013-09-03 == | ||
| - | * sequences arrived: [[:File: | + | * sequences arrived: [[:File:Heidelberg_PIK1.11-pIK1.13-pIK1.14-2013-09-03.zip|PIK1.11-pIK1.13-pIK1.14-2013-09-03.zip]], [[:File:Heidelberg_PIK1.11-2013-09-03 VF2.clustal.txt|PIK1.11-2013-09-03 VF2.clustal.txt]], [[:File:Heidelberg_PIK1.13-2013-09-03 VF2.clustal.txt|PIK1.13-2013-09-03 VF2.clustal.txt]], [[:File:Heidelberg_PIK1.14-2013-09-03 VF2.clustal.txt|PIK1.14-2013-09-03 VF2.clustal.txt]] |
* insertions leading to stop-codon in pIK1.11, pIK1.13; two bases of RBS missing in pIK1.14 | * insertions leading to stop-codon in pIK1.11, pIK1.13; two bases of RBS missing in pIK1.14 | ||
* check wiki of Cambridge 2007 team, who constructed the permeability device: [http://www.ccbi.cam.ac.uk/iGEM2007/index.php/Detailed_background_information_-_signalling#Plan they tried different promoter strengths], [http://www.ccbi.cam.ac.uk/iGEM2007/index.php/Signalling_project_description#Standard_Construction_of_Permeability_Device only the weakest promoter gave colonies] => bacteria might be selecting against the permeability device | * check wiki of Cambridge 2007 team, who constructed the permeability device: [http://www.ccbi.cam.ac.uk/iGEM2007/index.php/Detailed_background_information_-_signalling#Plan they tried different promoter strengths], [http://www.ccbi.cam.ac.uk/iGEM2007/index.php/Signalling_project_description#Standard_Construction_of_Permeability_Device only the weakest promoter gave colonies] => bacteria might be selecting against the permeability device | ||
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== 2013-09-04 == | == 2013-09-04 == | ||
| - | [[File: | + | [[File:Heidelberg_Methylmalonyl-digest2013-09-05.png|150px|thumb|left|Lane 1: NEB 2-log; lanes 2: pIK1.12 digested with BamHI+HindIII ]] |
| - | * sequences arrived: [[:File: | + | * sequences arrived: [[:File:Heidelberg_PIK1.14-2013-09-04.zip|PIK1.14-2013-09-04.zip]], [[:File:Heidelberg_PIK1.14-2013-09-04 IK25.clustal.txt|PIK1.14-2013-09-04 IK25.clustal.txt]], [[:File:Heidelberg_PIK1.14-2013-09-04 VR.clustal.txt|PIK1.14-2013-09-04 VR.clustal.txt]] |
* sequences match | * sequences match | ||
* [[Preparing glycerol stocks|prepare glycerol stock]] of DH10ß-pIK1.14 | * [[Preparing glycerol stocks|prepare glycerol stock]] of DH10ß-pIK1.14 | ||
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== 2013-09-05 == | == 2013-09-05 == | ||
| - | * sequence arrived: [[:File: | + | * sequence arrived: [[:File:Heidelberg_PIK1.12-2013-09-05.zip|PIK1.12-2013-09-05.zip]], [[:File:Heidelberg_PIK1.12-2013-09-05 VF2.clustal.txt|PIK1.12-2013-09-05 VF2.clustal.txt]] |
* ca. 1 kb missing from end of permeability device | * ca. 1 kb missing from end of permeability device | ||
Revision as of 23:39, 4 October 2013
Contents |
2013-09-02
- send pIK1.11, pIK1.13, pIK1.14 to sequencing with primer VF2
2013-09-03
- sequences arrived: PIK1.11-pIK1.13-pIK1.14-2013-09-03.zip, PIK1.11-2013-09-03 VF2.clustal.txt, PIK1.13-2013-09-03 VF2.clustal.txt, PIK1.14-2013-09-03 VF2.clustal.txt
- insertions leading to stop-codon in pIK1.11, pIK1.13; two bases of RBS missing in pIK1.14
- check wiki of Cambridge 2007 team, who constructed the permeability device: [http://www.ccbi.cam.ac.uk/iGEM2007/index.php/Detailed_background_information_-_signalling#Plan they tried different promoter strengths], [http://www.ccbi.cam.ac.uk/iGEM2007/index.php/Signalling_project_description#Standard_Construction_of_Permeability_Device only the weakest promoter gave colonies] => bacteria might be selecting against the permeability device
- put pIK1.12 in incubator again, maybe it has the right sequence and grows slowly
- pIK1.12 did grow, make miniPrep -> 67 ng/µl in 38 µl
- send pIK1.14 to sequencing with primers IK25, VR
2013-09-04
- sequences arrived: PIK1.14-2013-09-04.zip, PIK1.14-2013-09-04 IK25.clustal.txt, PIK1.14-2013-09-04 VR.clustal.txt
- sequences match
- prepare glycerol stock of DH10ß-pIK1.14
- digest pIK1.12 with BamHI+HindIII (20 µl total volume, 134 ng DNA [2 µl of miniPrep from 2013-09-03], 0.5 µl enzyme)
- digest looks about right, send to sequencing with primer VF2
2013-09-05
- sequence arrived: PIK1.12-2013-09-05.zip, PIK1.12-2013-09-05 VF2.clustal.txt
- ca. 1 kb missing from end of permeability device
