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Template:Kyoto/Notebook/Aug 20
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| + | [[File:Igku Aug20Electrophoresis3(DT・E＋X).jpg]]<br> | ||
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Latest revision as of 08:42, 24 September 2013
Contents |
Aug 20
Miniprep
| DNA | concentration[µg/µL] | 260/280 | 260/230 |
|---|---|---|---|
| 8/18 Ptet -1 | 220 | 1.69 | 1.87 |
| 8/18 Ptet -2 | 210 | 1.69 | 1.71 |
| 8/18 RBS-tetR-DT -1 | 236 | 1.63 | 1.69 |
| 8/18 RBS-tetR-DT -2 | 182 | 1.65 | 1.98 |
| 8/18 J23100-RBS-GFP-DT -1 | 334 | 1.71 | 2.12 |
| 8/18 J23100-RBS-luxR-DT -1 | 440 | 1.67 | 1.60 |
| 8/18 J23100-RBS-luxR-DT -2 | 344 | 1.68 | 1.83 |
| 8/18 J23100-RBS-lacZα-DT -1 | 280 | 1.70 | 1.81 |
| 8/18 RBS-lysis3 -1 | 282 | 1.67 | 1.76 |
| 8/18 RBS-lysis3 -2 | 212 | 1.30 | 1.35 |
LB Medium Plate
| volume | 200mL |
|---|---|
| Bacto Trypton | 2g |
| Bacto yeast extract | 1g |
| Nacl | 1g |
| Agar Powder | 2g |
| Amp | 40µl |
1. Measuring reagents and put they in the Erlenmeyer flask.
2. diluting in measuring cylinder to 200ml total.
3. autoclaving.
4. adding ampicirine 40ml.
Ristriction Enzyme Digestion
| 8/20 Ptet -1(220µg/mL) | EcoRI | XbaI | 10x BSA | 10x buffer M | MilliQ | total | |
|---|---|---|---|---|---|---|---|
| 2 cuts | 4.5 | 1 | 1 | 3 | 3 | 17.5 | 30 |
| 1 cut | 0.5 | 0.2 | 0 | 1 | 1 | 7.3 | 10 |
| 1 cut | 0.5 | 0 | 0.2 | 1 | 1 | 7.3 | 10 |
| NC | 0.5 | 0 | 0 | 1 | 1 | 7.5 | 10 |
| 8/20 RBS-tetR-DT -2(182µg/mL) | EcoRI | SpeI | 10x buffer H | MilliQ | total | |
|---|---|---|---|---|---|---|
| 2 cuts | 5.5 | 1 | 1 | 3 | 19.5 | 30 |
| 1 cut | 0.6 | 0.2 | 0 | 1 | 8.2 | 10 |
| 1 cut | 0.6 | 0 | 0.2 | 1 | 8.2 | 10 |
| NC | 0.6 | 0 | 0 | 1 | 8.4 | 10 |
| 8/20 J23100-RBS-GFP-DT(334µg/ml) | EcoRI | SpeI | 10x buffer H | MilliQ | total | |
|---|---|---|---|---|---|---|
| 2 cuts | 3.0 | 1 | 1 | 3 | 22 | 30 |
| 1 cut | 0.3 | 0.2 | 0 | 1 | 8.5 | 10 |
| 1 cut | 0.3 | 0 | 0.2 | 1 | 8.5 | 10 |
| NC | 0.3 | 0 | 0 | 1 | 8.7 | 10 |
| 8/20 J23100-RBS-luxR-DT -2(344µg/mL) | EcoRI | XbaI | BSA | 10x buffer M | MilliQ | total | |
|---|---|---|---|---|---|---|---|
| 2 cuts | 2.9 | 1 | 1 | 3 | 3 | 19.1 | 30 |
| 1 cut | 0.3 | 0.2 | 0 | 1 | 1 | 7.5 | 10 |
| 1 cut | 0.3 | 0 | 0.2 | 1 | 1 | 7.5 | 10 |
| NC | 0.3 | 0 | 0 | 1 | 1 | 7.7 | 10 |
| 8/20 RBS-lysis3 -1(282µg/mL) | EcoRI | SpeI | 10x buffer H | MilliQ | total | |
|---|---|---|---|---|---|---|
| 2 cuts | 3.5 | 1 | 1 | 3 | 21.5 | 30 |
| 1 cut | 0.4 | 0.2 | 0 | 1 | 8.4 | 10 |
| 1 cut | 0.4 | 0 | 0.2 | 1 | 8.4 | 10 |
| NC | 0.4 | 0 | 0 | 1 | 8.6 | 10 |
| 8/17 DT -1(188µg/mL) | EcoRI | XbaI | 10x BSA | 10x buffer M | MilliQ | total | |
|---|---|---|---|---|---|---|---|
| 2 cuts | 5.3 | 1 | 1 | 3 | 3 | 16.7 | 30 |
| 1 cut | 0.3 | 0.2 | 0 | 1 | 1 | 7.5 | 10 |
| 1 cut | 0.3 | 0 | 0.2 | 1 | 1 | 7.5 | 10 |
| NC | 0.3 | 0 | 0 | 1 | 1 | 7.7 | 10 |
Transformation
| Name | Sample | Competent Cells(XL10-gold) | Total | Plate |
|---|---|---|---|---|
| tRNA-Spinach-tRNA | 1µL | 10µL | 11µL | Amp |
| tetR-aptamer 12_PC(076) | 1µL | 10µL | 11µL | Amp |
| tetR-aptamer 12_1R(113) | 1µL | 10µL | 11µL | Amp |
| tetR-aptamer 12_1M(105) | 1µL | 10µL | 11µL | Amp |
| pT181 attenuator | 1µL | 10µL | 11µL | Amp |
| Fusin1 attenuator | 1µL | 10µL | 11µL | Amp |
| Fusion3m2 attenuator | 1µL | 10µL | 11µL | Amp |
| pT181 antisense | 1µL | 10µL | 11µL | Amp |
| Fusion1 antisense | 1µL | 10µL | 11µL | Amp |
| Fusion6 antisense | 1µL | 10µL | 11µL | Amp |
incubate 37°C overnight 20:34~
Electrophoresis
| Lane | Sample | Enzyme1 | Enzyme2 |
|---|---|---|---|
| 1 | 8/20 Ptet -1 | EcoRI | XbaI |
| 2 | 8/20 Ptet -1 | EcoRI | -- |
| 3 | 8/20 Ptet -1 | -- | XbaI |
| 4 | 8/20 Ptet -1 | -- | -- |
| 5 | 8/20 RBS-tetR-DT -2 | EcoRI | SpeI |
| 6 | 8/20 RBS-tetR-DT -2 | EcoRI | -- |
| 7 | 8/20 RBS-tetR-DT -2 | -- | SpeI |
| 8 | 8/20 RBS-tetR-DT -2 | -- | -- |
| 9 | 1kbp ladder | -- | -- |
| 10 | 8/20 J23100-RBS-GFP-DT -1 | EcoRI | SpeI |
| 11 | 8/20 J23100-RBS-GFP-DT -1 | EcoRI | -- |
| 12 | 8/20 J23100-RBS-GFP-DT -1 | -- | SpeI |
| 13 | 8/20 J23100-RBS-GFP-DT -1 | -- | -- |
| 14 | 8/20 J23100-RBS-luxR-DT -1 | EcoRI | XbaI |
| 15 | 8/20 J23100-RBS-luxR-DT -1 | EcoRI | -- |
| 16 | 8/20 J23100-RBS-luxR-DT -1 | -- | XbaI |
| 17 | 8/20 J23100-RBS-luxR-DT -1 | -- | -- |
| 18 | 1kbp ladder | -- | -- |
| 19 | 8/20 RBS-lysis3 -1 | EcoRI | SpeI |
| 20 | 8/20 RBS-lysis3 -1 | EcoRI | -- |
| 21 | 8/20 RBS-lysis3 -1 | -- | SpeI |
| 22 | 8/20 RBS-lysis3 -1 | -- | -- |
| 23 | 1kbp ladder | -- | -- |
| 24 | 8/17 DT | EcoRI | XbaI |
| 25 | 8/17 DT | EcoRI | -- |
| 26 | 8/17 DT | -- | XbaI |
| 27 | 8/17 DT | -- | -- |
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