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Template:Kyoto/Notebook/Aug 18
From 2013.igem.org
Contents |
Aug 18
Gel Extraction
| Lane | DNA | Enzyme |
|---|---|---|
| 1 | 100kb ladder | -- |
| 2 | 8/11 J23100 -② | -- |
| 3 | 8/11 J23100 -② | -- |
| 4 | -- | -- |
| 5 | 8/17 K117000(lysis1) -1 | -- |
| 6 | 8/17 K117000(lysis1) -1 | -- |
| 7 | -- | -- |
| 8 | 8/17 K748002(lysis3) -1 | -- |
| 9 | 8/17 K748002(lysis3) -1 | -- |
| 10 | -- | -- |
| 11 | 8/17 I13504(RBS-GFP-DT) -1 | -- |
| 12 | 8/17 I13504(RBS-GFP-DT) -1 | -- |
| 13 | 1kb ladder | -- |
| 14 | 8/17 I0462(RBS-luxR-DT) -1 | -- |
| 15 | 8/17 I0462(RBS-luxR-DT) -1 | -- |
| 16 | -- | -- |
| 17 | 8/17 I732020(RBS-lacZα-DT) -1 | -- |
| 18 | 8/17 I732020(RBS-lacZα-DT) -1 | -- |
| Name | concentration[µg/mL] | 260/280 | 260/230 |
|---|---|---|---|
| 8/14 J231000 | 1.7 | 1.61 | 0.04 |
| 8/17 K748002(lysis3) -1 | 5.4 | 2.39 | 0.45 |
| 8/17 I13504(RBS-GFP-DT) -1 | 3.4 | 3.33 | 0.19 |
| 8/17 I0462(RBS-luxR-DT) -1 | 4.4 | 2.07 | 0.37 |
| 8/17 I732020(RBS-lacZα-DT) -1 | 2.9 | 2.17 | 0.29 |
Ligation
| state | Vector | Inserter | Ligation High ver.2 | ||
|---|---|---|---|---|---|
| experiment | J23100 | 9.0 | RBS-lacZα-DT | 8.6 | |
| experiment | J23100 | 9.0 | RBS-GFP-DT | 7.3 | |
| experiment | RBS | 1 | lysis3 | 4.6 | |
| experiment | Plux | 1 | GFP | 7.3 | |
| experiment | J23100 | 9.0 | -- | -- | |
| experiment | RBS | 1.0 | -- | -- | |
| experiment | Plux | 1.0 | -- | -- | |
incubate 16°C 1hour
Restriction Enzyme Digestion
| 8/17K112806(lysis2)-1 | XbaI | PstI | Buffer | BSA | MilliQ | total | |
|---|---|---|---|---|---|---|---|
| 2 cuts | 2.0µL | 1.0µL | 1.0µL | 3.0µL | 3.0µL | 3.0µL | 30 µL |
| 1 cuts | 1.0&mic6/8ro;L | 0.2µL | -- | 1.0µL | 1.0µL | 6.8µL | 10 µL |
| 1 cuts | 1.0µL | -- | 0.2µL | 1.0µL | 1.0µL | 6.8µL | 10 µL |
Electrophoresis
| Lane | Sample | Enzyme1 | Enzyme2 |
|---|---|---|---|
| 1 | 100bp ladder | -- | -- |
| 2 | lysis1 | Xba1 | Pst1 |
| 3 | lysis1 | Xba1 | -- |
| 4 | lysis1 | -- | Pst1 |
| 5 | lysis1 | -- | -- |
| 6 | lysis2 | Xba1 | Pst1 |
| 7 | lysis2 | Xba1 | -- |
| 8 | lysis2 | -- | Pst1 |
| 9 | lysis2 | -- | -- |
| 10 | 100bp ladder | -- | -- |
Colony PCR
| Sample | base pair |
|---|---|
| 8/8 LBKana | -- |
| 8/16 Pbad araC | -- |
| negative control | -- |
| PreDenature | Denature | Annealing | Extension | cycle |
|---|---|---|---|---|
| 94°C | 94°C | 55°C | 68°C | -- |
| 5min | 30s | 30s | 1min |
Liquid Culture
| Sample | medium |
|---|---|
| 7/8 LB Kana | Plusgrow(+Kana) |
| 8/16 Pbad araC | Plusgrow(+Kana) |
Medium Plate
| volume | 200ml |
|---|---|
| Bacto Trypton | 2g |
| Bacto yeasy extract | 1g |
| NaCl | 1g |
| Agar Powder | 2g |
Transformation
| Name | Sample | Competent Cells | Total | Plate |
|---|---|---|---|---|
| 8/18 J23100+RBS-LacZα-DT | 2µL | 20µL | 22µL | Amp |
| 8/18 J23100+RBS-GFP-DT | 2µL | 20µL | 22µL | Amp |
| 8/18 J23100+RBS-luxR-DT | 2µL | 20µL | 22µL | Amp |
| RBS+lysis3 | 2µL | 20µL | 22µL | Amp |
| Plux+GFP | 2µL | 20µL | 22µL | CP |
| J23100 | 2µL | 20µL | 22µL | Amp |
| RBS | 2µL | 20µL | 22µL | Amp |
| Plux | 2µL | 20µL | 22µL | CP |
Electrophoresis
| Lane | Sample | Enzyme1 | Enzyme2 |
|---|---|---|---|
| 1 | 100bp ladder | -- | -- |
| 2 | 8/16Pbad araC-1 | -- | -- |
| 3 | 8/16Pbad araC-2 | -- | -- |
| 4 | negative control | -- | -- |
