Paris/July 25
From 2007.igem.org
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 |
