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Template:Kyoto/Notebook/Sep 6
From 2013.igem.org
Contents |
Sep 6
Electrophoresis
| Lane | Sample |
|---|---|
| 1 | 100bp ladder |
| 2 | 9/4 RBS-lysis3-DT -3 |
| 3 | 9/4 RBS-lysis3-DT -4 |
| 4 | 9/4 RBS-lysis3-DT -5 |
| 5 | 9/4 RBS-lysis1-DT -3 |
| 6 | 9/4 Ptrc-KaiC -1 |
| 7 | 9/4 pSB4K5 -1 |
| 8 | 100bp ladder |
Colony PCR
| Sample | base pair |
|---|---|
| Pconst-spinach-DT-(1~4) | 563 |
| Plac-spinach-DT-(1~4) | 659 |
| Pcon-pT181 antisense-(1~4) | 468 |
| Ptet-pT181 antisense-(1~4) | 604 |
| Pconst-pT181 attenuater-(1~4) | 568 |
| Pconst+RBS-tetR-DT-(1~4) | 1121 |
| Ptet+RBS-lacZα-DT-(1~4) | 1162 |
| PreDenature | Denature | Annealing | Extension | cycle |
|---|---|---|---|---|
| 94°C | 94°C | 55°C | 68°C | -- |
| 5min | 30s | 30s | 40s | 30cycles |
Liquid Culture
| Sample | medium |
|---|---|
| 9/4 RBS-Lysis3+DT3 | Plusgrow medium(+CP) |
| 9/4 RBS-Lysis1+DT3 | Plusgrow medium(+CP) |
| 9/4 pSB4K5 | Plusgrow medium(+Kan) |
Electrophoresis
| Lane | Sample |
|---|---|
| 1 | 100bp ladder |
| 2 | Pconst-spinach-DT -(1) |
| 3 | Pconst-spinach-DT -(2) |
| 4 | Pconst-spinach-DT -(3) |
| 5 | Pconst-spinach-DT -(4) |
| 6 | Plac-spinach-DT -(1) |
| 7 | Plac-spinach-DT -(2) |
| 8 | Plac-spinach-DT -(3) |
| 9 | Plac-spinach-DT -(4) |
| 10 | Pcon-pT181 antisense -(1) |
| 11 | Pcon-pT181 antisense -(2) |
| 12 | Pcon-pT181 antisense -(3) |
| 13 | Pcon-pT181 antisense -(4) |
| Lane | Sample |
|---|---|
| 1 | 100bp ladder |
| 2 | Ptet-pT181 antisense -(1) |
| 3 | Ptet-pT181 antisense -(2) |
| 4 | Ptet-pT181 antisense -(3) |
| 5 | Ptet-pT181 antisense -(4) |
| 6 | Plac-pT181 attenuater -(1) |
| 7 | Plac-pT181 attenuater -(2) |
| 8 | Plac-pT181 attenuater -(3) |
| 9 | Plac-pT181 attenuater -(4) |
Miniprep
| DNA | concentration [µg/mL] | 260/280 | 260/230 |
|---|---|---|---|
| pT181 antisense-(1) | 238.7 | 1.68 | 1.44 |
| pT181 antisense-(2) | 248.8 | 1.83 | 1.77 |
| pT181 antisense-(3) | 223.2 | 1.78 | 1.61 |
| pT181 antisense-(4) | 220.8 | 1.89 | 1.92 |
| tetR aptamer-(1) | 252.5 | 1.89 | 1.90 |
| tetR aptamer-(2) | 211.2 | 1.90 | 1.99 |
| DT181 attenuater | 102.2 | 1.98 | 2.17 |
| 121R aptamer-DT | 251.7 | 1.92 | 2.01 |
| pSB4K5 | 212.9 | 1.80 | 1.90 |
| RBS-lacZα-DT | 185.7 | 1.78 | 1.79 |
| RBS-lysis2-DT-(1) | 84.8 | 1.85 | 2.24 |
Restriction Enzyme Digestion
| 9/6 RBS-lysis2-DT (78.3 µg/mL) | XbaI | PstI | BSA | Buffer | MilliQ | total | |
|---|---|---|---|---|---|---|---|
| 2 cuts | 22 µL | 1 µL | 1 µL | 3 µL | 3 µL | 0 µL | 30 µL |
| NC | 1.3 µL | 0 µL | 0 µL | 1 µL | 1 µL | 6.7 µL | 10 µL |
| aptamer-DT (251.7 µg/mL) | XbaI | PstI | BSA | Buffer | MilliQ | total | |
|---|---|---|---|---|---|---|---|
| 2 cuts | 0.4 µL | 1 µL | 1 µL | 3 µL | 3 µL | 14.1 µL | 30 µL |
| NC | 0.4 µL | 0 µL | 0 µL | 1 µL | 1 µL | 7.6 µL | 10 µL |
| Pλ-luxI (86.4 µg/mL) | EcoRI | SpeI | BSA | Buffer | MilliQ | total | |
|---|---|---|---|---|---|---|---|
| 2 cuts | 23.1 µL | 1 µL | 1 µL | 3 µL | 3 µL | 1.9 µL | 30 µL |
| NC | 1.2 µL | 0 µL | 0 µL | 1 µL | 1 µL | 6.8 µL | 10 µL |
- at 37 °C, for 1h
Colony PCR
| Sample | base pair |
|---|---|
| Pconst+RBS-tetR-DT-(1) | 1121 |
| Pconst+RBS-tetR-DT-(2) | 1121 |
| Pconst+RBS-tetR-DT-(3) | 1121 |
| Pconst+RBS-tetR-DT-(4) | 1121 |
| Ptet+RBS-lacZα-DT-(1) | 1162 |
| Ptet+RBS-lacZα-DT-(2) | 1162 |
| Ptet+RBS-lacZα-DT-(3) | 1162 |
| Ptet+RBS-lacZα-DT-(4) | 1162 |
Gel Extraction
| Lane | DNA | Enzyme |
|---|---|---|
| 1 | 100bp ladder | -- |
| 2 | RBS-lysis2-DT | XbaI & PstI |
| 3 | ||
| 4 | -- | -- |
| 5 | aptamer 12_1R-DT | XbaI & PstI |
| 6 |
| Lane | DNA | Enzyme |
|---|---|---|
| 1 | 100bp ladder | -- |
| 2 | Pλ-luxI | EcoRI & SpeI |
| 3 |
| Name | concentration[µg/mL] | 260/280 | 260/230 |
|---|---|---|---|
| RBS-lysis2-DT (XbaI & PstI) | 7.1 | 1.88 | 0.07 |
| aptamer 12_1R-DT (XbaI & PstI) | 15.3 | 1.93 | 0.70 |
| Pλ-luxI (EcoRI & SpeI) | 4.5 | 1.79 | 0.31 |
Liquid Culture
| Sample | medium |
|---|---|
| 9/5 Pconst-spinach-DT-(1) | Plusgrow medium(+Amp&CP) |
| 9/5 Ptet-pT181 antisense-(1) | Plusgrow medium(+CP) |
| 9/5 Pconst+RBS-tetR-DT-(1) | Plusgrow medium(+Amp&CP) |
| 9/5 Ptet+RBS-lacZα-DT-(1) | Plusgrow medium(+CP) |
| 9/5 Pcon-pT181 attenuater-(1) | Plusgrow medium(+CP) |
| 9/4 pT181 antisense-(1) | Plusgrow medium(+CP) |
| 9/4 aptamer 12-1R-(1) | Plusgrow medium(+CP) |
Colony PCR
| Sample | base pair |
|---|---|
| 9/5 Plac-spinach-DT-(5~12) | 563 |
| 9/5 Pcon-antisense-(5~12) | 468 |
| 9/5 Plac-attenuater-(5~12) | 664 |
| 9/5 pT181 antisense+Plac-(1) | - |
| PreDenature | Denature | Annealing | Extension | cycle |
|---|---|---|---|---|
| 94°C | 94°C | 55°C | 68°C | -- |
| 5min | 30s | 30s | 42s | 30cycles |
