From 2013.igem.org
Sep 13
Miniprep
Nakamoto
DNA | concentration[µg/mL] | 260/280 | 260/230
|
9/12 Fusion6 antisense | 245.9 | 1.88 | 1.41
|
9/12 Plux-3 | 115.5 | 1.58 | 1.32
|
9/12 Plux-4 | 125.3 | 1.93 | 1.44
|
9/12 Fusion3in2 attenuator-1 | 231.9 | 1.81 | 1.31
|
Lane | DNA | Enzyme
|
1 | 1kb ladder | --
|
3 | RBS-lysis2-DT | XbaI&PstI
|
4 | RBS-lysis2-DT | XbaI&PstI
|
5 | RBS-lysis2-DT | XbaI&PstI
|
File:Igku Aug19electrophoresis3
File:Igku Aug19electrophoresis4
Name | concentration[µg/mL] | 260/280 | 260/230
|
RBS-lacZα-DT(EcoRI&SpeI) | 9.8 | 1.50 | 0.39
|
Pcon-RBS-lacZα-DT(EcoRI&SpeI) | 11.2 | 1.47 | 0.08
|
</div>
Lane | DNA | Enzyme
|
3 | RBS-lysis3-DT | XbaI&PstI
|
4 | RBS-lysis3-DT | XbaI&PstI
|
5 | RBS-lysis3-DT | XbaI&PstI
|
6 | 1kb ladder | --
|
File:Igku Aug19electrophoresis3
File:Igku Aug19electrophoresis4
Name | concentration[µg/mL] | 260/280 | 260/230
|
RBS-lacZα-DT(EcoRI&SpeI) | 9.8 | 1.50 | 0.39
|
Pcon-RBS-lacZα-DT(EcoRI&SpeI) | 11.2 | 1.47 | 0.08
|
</div>
Lane | DNA | Enzyme
|
2 | 1kb ladder | --
|
3 | pSB1C3 | EcoRI&SpeI
|
4 | pSB1C3 | EcoRI&SpeI
|
5 | pSB1C3 | EcoRI&SpeI
|
File:Igku Aug19electrophoresis3
File:Igku Aug19electrophoresis4
Name | concentration[µg/mL] | 260/280 | 260/230
|
RBS-lacZα-DT(EcoRI&SpeI) | 9.8 | 1.50 | 0.39
|
Pcon-RBS-lacZα-DT(EcoRI&SpeI) | 11.2 | 1.47 | 0.08
|
</div>
Lane | DNA | Enzyme
|
1 | 1kb ladder | --
|
2 | pSB1C3 | XbaI&PstI
|
3 | pSB1C3 | XbaI&PstI
|
4 | pSB1C3 | XbaI&PstI
|
File:Igku Aug19electrophoresis3
File:Igku Aug19electrophoresis4
Name | concentration[µg/mL] | 260/280 | 260/230
|
RBS-lacZα-DT(EcoRI&SpeI) | 9.8 | 1.50 | 0.39
|
Pcon-RBS-lacZα-DT(EcoRI&SpeI) | 11.2 | 1.47 | 0.08
|
</div>
Restriction Enzyme Digestion
Nakamoto
| 9/13 Plac | EcoRI | SpeI | XbaI | PstI | Buffer | BSA | MilliQ | total
|
1cuts(PstI) | 16 | 0µL | 0µL | 0µL | 1µL | 3µL | 3µL | 7µL | 30µL
|
NC | 0.8 | 0µL | 0µL | 0µL | 0µL | 1µL | 1µL | 7.2µL | 10µL
|
| 9/10 Pcon attenuator | EcoRI | SpeI | XbaI | PstI | Buffer | BSA | MilliQ | total
|
1cut(PstI) | 10 | 0µL | 0µL | 0µL | 1µL | 3µL | 3µL | 13µL | 30µL
|
NC | 0.5 | 0µL | | 0µL | 0µL | 0µL | 1µL | 1µL | 7.5µL | 10µL
|
| 9/12 Plux | EcoRI | SpeI | XbaI | PstI | Buffer | BSA | MilliQ | total
|
1cuts(PstI) | 12 | 0µL | 0µL | 0µL | 1µL | 3µL | 3µL | 11µL | 30µL
|
NC | 0.6 | 0µL | 0µL | 0µL | 0µL | 1µL | 1µL | 7.4µL | 10µL
|
| 9/12 DT-E | EcoRI | SpeI | XbaI | PstI | Buffer | BSA | MilliQ | total
|
1cut(XbaI) | 23.7 | 0µL | 0µL | 1µL | 0µL | 3µL | 3µL | 0µL | 30.7µL
|
NC | 1.2 | 0µL | 0µL | 0µL | 0µL | 1µL | 1µL | 6.8µL | 10µL
|
| 9/12 Pcon-RBS-luxR-bTE | EcoRI | SpeI | XbaI | PstI | Buffer | BSA | MilliQ | total
|
1cuts(XbaI) | 22.4 | 0µL | 0µL | 1µL | 0µL | 3µL | 3µL | 0.6µL | 30µL
|
NC | 2.2 | 0µL | 0µL | 0µL | 0µL | 1µL | 1µL | 5.8µL | 10µL
|
| 9/12 Pcon-P | EcoRI | SpeI | XbaI | PstI | Buffer | BSA | MilliQ | total
|
1cut(SpeI) | 21 | 0µL | 1µL | 0µL | 0µL | 3µL | 3µL | 12.0µL | 30µL
|
NC | 2.1 | 0µL | 0µL | 0µL | 0µL | 1µL | 1µL | 5.7µL | 10µL
|
M9 (EDTA)
volume | 10ml
|
Na2HPO4 | 300mg
|
KH2PO4 | 150mg
|
NaCl | 25mg
|
NH4Cl | 50mg
|
Fe(3)-EDTA | 21mg
|
5mM EDTA</div>
volume | 10ml
|
Na2HPO4 | 300mg
|
KH2PO4 | 150mg
|
NaCl | 25mg
|
NH4Cl | 50mg
|
Fe(3)-EDTA | 105mg
|
</div>
Liquid Culture
Tatsui
Sample | medium
|
9/11 aptamer12_1M(pSB1C3)-1 | --
|
9/11 aptamer 12_P1 | --
|
9/11 Fusaion1 antisense-4 | --
|
9/12 pSB4K5-1 | --
|
9/12 pSB4K5-2 | --
|
Restriction Enzyme Digestion
Nakamoto
| 9/13 Plac-P | EcoRI | SpeI | XbaI | PstI | Buffer | BSA | MilliQ | total
|
1cuts(SpeI) | 23.5 | 0µL | 0.5µL | 0µL | 0µL | 3µL | 3µL | 0µL | 30µL
|
NC | 8 | 0µL | 0µL | 0µL | 0µL | 1µL | 1µL | 0µL | 10µL
|
| 9/13 Pcon-pT181 attenuator-P | EcoRI | SpeI | XbaI | PstI | Buffer | BSA | MilliQ | total
|
1cut(SpeI) | 23.5 | 0µL | 0.5µL | 0µL | 0µL | 3µL | 3µL | 0µL | 30µL
|
NC | 02.3 | 0µL | | 0µL | 0µL | 0µL | 1µL | 1µL | 5.7µL | 10µL
|
| 9/13 Plux-P | EcoRI | SpeI | XbaI | PstI | Buffer | BSA | MilliQ | total
|
1cuts(SpeI) | 23.8 | 0µL | 0.2µL | 0µL | 1µL | 3µL | 3µL | 0µL | 30µL
|
NC | 8 | 0µL | 0µL | 0µL | 0µL | 1µL | 1µL | 0µL | 10µL
|
| 9/5 RBS-lacZα-DT | EcoRI | SpeI | XbaI | PstI | Buffer | BSA | MilliQ | total
|
2cut(XbaI&PstI) | 10.8 | 0µL | 0µL | 1µL | 1µL | 3µL | 3µL | 11.2µL | 30µL
|
NC | 0.5 | 0µL | 0µL | 0µL | 0µL | 1µL | 1µL | 7.5µL | 10µL
|
Electrophoresis
Ligation
No Name
state | Vector | Inserter | Ligation High ver.2
|
experiment | 9/13 Plac (SpeI&PstI) | 8.5µL | 9/13 pT181 antisense (XbaI&PstI) | 2.1 µL | 10.6 µL
|
experiment | 9/13 Plac (SpeI&PstI) | 8.5 µL | 9/10 aptamer 12_1R-DT (XbaI&PstI) | 1.8 µL | 10 .3µL
|
experiment | 9/13 Plac (SpeI&PstI) | 8.5 µL | 9/2 pT181 attenuator(XbaI&PstI) | 2.3 µL | 10.8 µL
|
experiment | 9/13 Plux (SpeI&PstI) | 11.4 µL | 9/13 RBS-lysis1-DT (XbaI&PstI) | 5.0 µL | 16.4 µL
|
experiment | 9/13 Plux (SpeI&PstI) | 11.4 µL | 9/13 RBS-lysis2-DT (XbaI&PstI) | 7.0 µL | 18.4 µL
|
experiment | 9/13 Plux (SpeI&PstI) | 7.2 µL | 9/13 RBS-lysis3-DT (XbaI&PstI) | 6.0 µL | 13.2 µL
|
experiment | 9/13 Pcon-RBS-luxR-DT (EcoRI&XbaI) | 2.0 µL | 9/4 Plux-RBS-GFP-DT (EcoRI&SpeI) | 5.2 µL | 72 µL
|
experiment | 9/13 Pcon-RBS-luxR-DT (EcoRI&XbaI) | 2.0 µL | 9/10 aptamer 12_1R-DT (XbaI&PstI) | 1.6 µL | 3.6 µL
|
experiment | 9/12 Ptet-pT181 antisense (--) | 1.0 µL | 9/10 spinach-DT (--) | 2.4 µL | 3.4 µL
|
PCR
No name
genome DNA | KOD plus | 10x buffer | dNTP | MgSO4 | SasA_fwd primer | SasA_rev primer | MilliQ | total
|
| 0.5 | 2.5 | 2.5 | 1.5 | 0.75 | 0.75 | | 25
|
PreDenature | Denature | Annealing | Extension | cycle
|
°C | °C | °C | °C | --
|
(◎Д◎) | | | |
|
PCR
No name
genome DNA | KOD plus | 10x buffer | dNTP | MgSO4 | SasA_fwd primer | SasA_rev primer | MilliQ | total
|
| 0.5 | 2.5 | 2.5 | 1.5 | 0.75 | 0.75 | | 25
|
PreDenature | Denature | Annealing | Extension | cycle
|
°C | °C | °C | °C | --
|
(◎Д◎) | | | |
|
Transformation
No name
Name | Sample | Competent Cells | Total | Plate
|
9/13 Plac+pT181 antisense | 2µL | 20µL | 22µL | Amp
|
9/13 Plac+apt12_1R | 2µL | 20µL | 22µL | Amp
|
9/13 Plac+pT181 attenuator | 2µL | 20µL | 22µL | Amp
|
9/13 Plux+pT181 antisense | 2µL | 20µL | 22µL | CP
|
9/13 Plux+RBS-lysis2-DT | 2µL | 20µL | 22µL | CP
|
9/13 Plux+RBS-lysis2-DT | 2µL | 20µL | 22µL | CP
|
9/13 Pcon-RBS-lacR-DT+Plux-RBS-GFP-DT | 2µL | 20µL | 22µL | Amp
|
9/13 Pcon-pT181 antisense+apt12_1R-DT | 2µL | 20µL | 22µL | Amp
|
9/13 Ptet-pT181antisense+spinach-DT | 2µL | 20µL | 22µL | CP
|
Pλ-luxI-1 | 2µL | 20µL | 22µL | Amp
|