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Team:Freiburg/Notebook
From 2012.igem.org
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{{#calendar: title=Freiburg |year=2012 | month=09}} | {{#calendar: title=Freiburg |year=2012 | month=09}} | ||
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| + | week 06/04/12 - 06/10/12 | ||
| + | • Golden-Gate-Cloning-Reaction (GGC) to assemble a 12-repeat-TAL-TF, that recognizes the sequence TACATTGGACCTAT --> protocol: sanjana et al. (first hexamers...) | ||
| + | • transformation: GGC-reaction | ||
| + | • making aliquots of ordered GGC-Primers (freiGEM-method) | ||
| + | • making aliquots of ordered, i.e. synthesized, direpeats | ||
| + | • extension-PCR of direpeats with ordered freiGEM-GGC- | ||
| + | • PCR-Purification of extension-PCR | ||
| + | • mutagenesis-PCR of Zhang-plasmid to remove restriction sites (SpeI, PstI, EcoRI, XbaI) --> making the plasmid work with iGEM-standard | ||
| + | week 06/11/12 - 06/16/12 | ||
| + | • optimizing PCR conditions for extension of direpeats | ||
| + | • redoing GGC-reaction --> protocol: sanjana et al. | ||
| + | • transformation of redone GGC-reaction | ||
| + | • GGC á la freiGEM --> transformation | ||
| + | • testing of iGEM-distribution kit | ||
| + | week 06/17/12 - 06/23/12 | ||
| + | • cloning of direpeats into pJET 1.2 vector-system | ||
| + | • colony-PCR of freiGEM-GGC product | ||
| + | • place sequencing order for freiGEM-GGC | ||
| + | week 06/24/12 - 07/01/12 | ||
| + | • optimizing of freiGEM-GGC under various conditions | ||
| + | • transformation of pJET-direpeats into bacteria | ||
| + | • using the iGEM distribution kit: Bba_A12564 (CMV+Luc) --> reporter-plasmid for DNA-methyl-transferase testing | ||
| + | • Miniprep of GGC-transformation (1 successful) | ||
| + | week 07/02/12 - 07/08/12 | ||
| + | • extension-PCR with all 96 direpeats on one well-plate | ||
| + | • transformation | ||
| + | • PCR-amplification of all 4 iGEM-backbones --> testing different conditions | ||
| + | • making bacteria competent for transformation | ||
| + | week 07/09/12 - 07/15/12 | ||
| + | • digest of exDirepeats with XbaI and PstI | ||
| + | • nanodrop of exDirepeats | ||
| + | • ligation of exDirepeats in psB1C3 vector backbone and then transformation | ||
| + | • colony-PCR, gel run, making cultures | ||
| + | • miniprep of some of the 96 exDirepeats | ||
| + | week 07/16/12 - 07/22/12 | ||
| + | • Transformation of synthesis products (Dnmt, hyperactive Gin, Gin L7C7, Tn3 and TAL-ORF) into DH10B-strain competent cells | ||
| + | • amplification of psb1c3 vector backbone, then gel-run and gel-purification | ||
| + | • picking of colonies (transformation of synthesis-products) | ||
| + | • miniprep of synthesis-products | ||
| + | • repeat of pcr-amplification of psb1c3 vector backbone | ||
| + | week 23/07/12 - 07/29/12 | ||
| + | week 07/30/12 - 08/05/12 | ||
| + | • GGC to produce freiGEM-Mammo-Brick with CMV-promotor, Puromycin-resistance and postORF and TALEN-NG-vector backbone | ||
| + | • to do so a extension-PCR on the parts was done | ||
| + | • CMV-Promotor was taken out of iGEM Distribution Kit 2012 | ||
| + | week 08/06/12 - 08/12/12 | ||
| + | • mutagenesis-PCR of pcb1c3 vector backbone to eliminate restriction site for BsmBI | ||
| + | • repeat of GGC to get MammoBrick | ||
| + | • gel-run of mutagenesis-PCR of psb1c3 | ||
| + | week 08/13/12 - 08/19/12 | ||
| + | |||
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|[[Image:smallbarrev.png|960px|no frame]] | |[[Image:smallbarrev.png|960px|no frame]] | ||
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Revision as of 22:03, 15 September 2012
Notebook
| September | ||||||
| M | T | W | T | F | S | S |
| 1 | 2 | |||||
| 3 | 4 | 5 | 6 | 7 | 8 | 9 |
| 10 | 11 | 12 | 13 | 14 | 15 | 16 |
| 17 | 18 | 19 | 20 | 21 | 22 | 23 |
| 24 | 25 | 26 | 27 | 28 | 29 | 30 |
week 06/04/12 - 06/10/12
• Golden-Gate-Cloning-Reaction (GGC) to assemble a 12-repeat-TAL-TF, that recognizes the sequence TACATTGGACCTAT --> protocol: sanjana et al. (first hexamers...) • transformation: GGC-reaction • making aliquots of ordered GGC-Primers (freiGEM-method) • making aliquots of ordered, i.e. synthesized, direpeats • extension-PCR of direpeats with ordered freiGEM-GGC- • PCR-Purification of extension-PCR • mutagenesis-PCR of Zhang-plasmid to remove restriction sites (SpeI, PstI, EcoRI, XbaI) --> making the plasmid work with iGEM-standard
week 06/11/12 - 06/16/12
• optimizing PCR conditions for extension of direpeats • redoing GGC-reaction --> protocol: sanjana et al. • transformation of redone GGC-reaction • GGC á la freiGEM --> transformation • testing of iGEM-distribution kit
week 06/17/12 - 06/23/12
• cloning of direpeats into pJET 1.2 vector-system • colony-PCR of freiGEM-GGC product • place sequencing order for freiGEM-GGC
week 06/24/12 - 07/01/12
• optimizing of freiGEM-GGC under various conditions • transformation of pJET-direpeats into bacteria • using the iGEM distribution kit: Bba_A12564 (CMV+Luc) --> reporter-plasmid for DNA-methyl-transferase testing • Miniprep of GGC-transformation (1 successful)
week 07/02/12 - 07/08/12
• extension-PCR with all 96 direpeats on one well-plate • transformation • PCR-amplification of all 4 iGEM-backbones --> testing different conditions • making bacteria competent for transformation
week 07/09/12 - 07/15/12
• digest of exDirepeats with XbaI and PstI • nanodrop of exDirepeats • ligation of exDirepeats in psB1C3 vector backbone and then transformation • colony-PCR, gel run, making cultures • miniprep of some of the 96 exDirepeats
week 07/16/12 - 07/22/12
• Transformation of synthesis products (Dnmt, hyperactive Gin, Gin L7C7, Tn3 and TAL-ORF) into DH10B-strain competent cells • amplification of psb1c3 vector backbone, then gel-run and gel-purification • picking of colonies (transformation of synthesis-products) • miniprep of synthesis-products • repeat of pcr-amplification of psb1c3 vector backbone
week 23/07/12 - 07/29/12 week 07/30/12 - 08/05/12
• GGC to produce freiGEM-Mammo-Brick with CMV-promotor, Puromycin-resistance and postORF and TALEN-NG-vector backbone • to do so a extension-PCR on the parts was done • CMV-Promotor was taken out of iGEM Distribution Kit 2012
week 08/06/12 - 08/12/12
• mutagenesis-PCR of pcb1c3 vector backbone to eliminate restriction site for BsmBI • repeat of GGC to get MammoBrick • gel-run of mutagenesis-PCR of psb1c3
week 08/13/12 - 08/19/12
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