Paris/July 25
From 2007.igem.org
(Difference between revisions)
Eismoustique (Talk | contribs) (→Ligation & transformation reactions) |
Eismoustique (Talk | contribs) (→Ligation & transformation reactions) |
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| Line 90: | Line 90: | ||
|D9.1 (J61002 ready for insertion of a FI) | |D9.1 (J61002 ready for insertion of a FI) | ||
|1.5 | |1.5 | ||
| - | | | + | |construct with pBad promotor regulating expression of mRFP (in order to test activity of pBad) |
|- style="background: #cccccc;" | |- style="background: #cccccc;" | ||
| style="background: #ccffcc;" |L2 | | style="background: #ccffcc;" |L2 | ||
| Line 97: | Line 97: | ||
|D22.1 (pSB1A2 Eco, Pst) | |D22.1 (pSB1A2 Eco, Pst) | ||
|3 | |3 | ||
| - | | | + | |Insertion of lox66DapAE.coli construct as a biobrick in pSB1A2 plasmid |
|- style="background: #cccccc;" | |- style="background: #cccccc;" | ||
| style="background: #ccffcc;" |L3 | | style="background: #ccffcc;" |L3 | ||
| Line 104: | Line 104: | ||
|D22.1 (pSB1A2 Eco, Pst) | |D22.1 (pSB1A2 Eco, Pst) | ||
|3 | |3 | ||
| - | | | + | |Insertion of lox66DapAsubtilis construct as a biobrick in pSB1A2 plasmid |
|- style="background: #cccccc;" | |- style="background: #cccccc;" | ||
| style="background: #ccffcc;" |L4 | | style="background: #ccffcc;" |L4 | ||
| Line 111: | Line 111: | ||
|D1 (pJ23100 BV) | |D1 (pJ23100 BV) | ||
|2 | |2 | ||
| - | | | + | |lo66-DapAE.coli under regulation of strong promotor pJ23100 |
|- style="background: #cccccc;" | |- style="background: #cccccc;" | ||
| style="background: #ccffcc;" |L5 | | style="background: #ccffcc;" |L5 | ||
| Line 118: | Line 118: | ||
|D1 (pJ23100 BV) | |D1 (pJ23100 BV) | ||
|2 (+1.5µL H2O) | |2 (+1.5µL H2O) | ||
| - | | | + | |lox66-DapAsubtilis under regulation of strong promotor pJ23100 |
|- style="background: #cccccc;" | |- style="background: #cccccc;" | ||
| style="background: #ccffcc;" |L6 | | style="background: #ccffcc;" |L6 | ||
| Line 125: | Line 125: | ||
|D15.1 (B0030 BV) | |D15.1 (B0030 BV) | ||
|2 | |2 | ||
| - | | | + | |construction of the biobrick strong RBS(B0030)-Cre ORF |
|- style="background: #cccccc;" | |- style="background: #cccccc;" | ||
| style="background: #ccffcc;" |L7 | | style="background: #ccffcc;" |L7 | ||
| Line 132: | Line 132: | ||
|D1 (pJ23100 BV) | |D1 (pJ23100 BV) | ||
|2 (+2µL H2O) | |2 (+2µL H2O) | ||
| - | | | + | |lox71-ftsZ under regulation of strong promotor pJ23100 |
|- style="background: #cccccc;" | |- style="background: #cccccc;" | ||
| style="background: #ccffcc;" |L8 | | style="background: #ccffcc;" |L8 | ||
| Line 139: | Line 139: | ||
|D3 (pJ23107 BV) | |D3 (pJ23107 BV) | ||
|2 (+2 H2O) | |2 (+2 H2O) | ||
| - | | | + | |lox71-ftsZ under regulation of promotor pJ23107 (intermediate strength) |
|- style="background: #cccccc;" | |- style="background: #cccccc;" | ||
| style="background: #ccffcc;" |L9 | | style="background: #ccffcc;" |L9 | ||
| Line 146: | Line 146: | ||
|D22.1 (pSB1A2 Eco, Pst) | |D22.1 (pSB1A2 Eco, Pst) | ||
|2 (+6 H2O) | |2 (+6 H2O) | ||
| - | | | + | |lox66 cloned as a biobrick in pSB1A2 |
|- style="background: #cccccc;" | |- style="background: #cccccc;" | ||
| style="background: #ccffcc;" |L10 | | style="background: #ccffcc;" |L10 | ||
| Line 153: | Line 153: | ||
|D22.1 (pSB1A2 Eco, Pst) | |D22.1 (pSB1A2 Eco, Pst) | ||
|2 (+6 H2O) | |2 (+6 H2O) | ||
| - | | | + | |lox71 cloned as a biobrick |
|} | |} | ||
Revision as of 11:11, 27 July 2007
Digestions
Digestion of purifications of PCR products
| Digestion Products | ||||||||
|---|---|---|---|---|---|---|---|---|
| Number | Product Name | Matrix Name | Enzyme 1 | Enzyme 2 | size | Description | ||
| D28 | RBS-DapAColi BB | P8 (RBS-DapAColi) | EcoRI | PstI | ~950 | Biobrick | ||
| D29 | RBS-DapAColi | P8 (RBS-DapAColi) | XbaI | PstI | ~950 | Backward insert | ||
| D30 | RBS-DapASubtilis BB | P9 (RBS-DapASubtilis) | EcoRI | PstI | ~950 | Biobrick | ||
| D31 | RBS-DapASubtilis | P9 (RBS-DapASubtilis) | XbaI | PstI | ~950 | Backward insert | ||
| D32 | Lox71-FtsA-FtsZ | P10 Lox71-FtsA-FtsZ | XbaI | PstI | ~2600 | Backward insert - EcoRI site still presents : we can't use EcoRI | ||
Ligation & transformation reactions
The Ligation reactions performed yesterday have all failed (the reactions were performed at 42° by mistake, then, another 1µL of ligase was added for a 5min rxn at 25°)
The following reactions were performed today
| Ligations | |||||
|---|---|---|---|---|---|
| Number | Insert | Insert Volume (µL) | Vector | Vector Volume (µL) | Comments |
| L1 | D23.1 (AraC/pBad promoter FI) | 8.5 | D9.1 (J61002 ready for insertion of a FI) | 1.5 | construct with pBad promotor regulating expression of mRFP (in order to test activity of pBad) |
| L2 | D18 (BB dig. lox66DapAE.coli) | 7 | D22.1 (pSB1A2 Eco, Pst) | 3 | Insertion of lox66DapAE.coli construct as a biobrick in pSB1A2 plasmid |
| L3 | D19 (BB dig. lox66DapAsubtilis) | 7 | D22.1 (pSB1A2 Eco, Pst) | 3 | Insertion of lox66DapAsubtilis construct as a biobrick in pSB1A2 plasmid |
| L4 | D20 (Lox66-DapAE.coli BI) | 8 | D1 (pJ23100 BV) | 2 | lo66-DapAE.coli under regulation of strong promotor pJ23100 |
| L5 | D21 (lox66-DapAsubtilis BI) | 6.5 | D1 (pJ23100 BV) | 2 (+1.5µL H2O) | lox66-DapAsubtilis under regulation of strong promotor pJ23100 |
| L6 | D14.1 (Cre ORF) | 8 | D15.1 (B0030 BV) | 2 | construction of the biobrick strong RBS(B0030)-Cre ORF |
| L7 | D27 (Lox71-ftsZ BI) | 6 | D1 (pJ23100 BV) | 2 (+2µL H2O) | lox71-ftsZ under regulation of strong promotor pJ23100 |
| L8 | D27 (Lox71-ftsZ BI) | 6 | D3 (pJ23107 BV) | 2 (+2 H2O) | lox71-ftsZ under regulation of promotor pJ23107 (intermediate strength) |
| L9 | lox66 [O14+O15] (0.2µM) | 2 | D22.1 (pSB1A2 Eco, Pst) | 2 (+6 H2O) | lox66 cloned as a biobrick in pSB1A2 |
| L10 | lox71 [O16+O17] (0.2µM) | 2 | D22.1 (pSB1A2 Eco, Pst) | 2 (+6 H2O) | lox71 cloned as a biobrick |
