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Team:Chiba/jk/γ
From 2008.igem.org
17,August 2008
PCR
BBa_E0430①
BBa_E0420②
BBa_J06702③
BBa_S01416④
BBa_I73003⑤
BBa_I73004⑥
BBa_I730026⑦
BBa_I763002⑧
| Sample No | ① | ② | ③ | ④ | ⑤ | ⑥ | ⑦ | ⑧ |
| DNA tamplate | 1 | 1 | 1 | 1 | >1 | 1 | 1 | 1 |
| FW primer | 5 | 5 | 5 | 5 | 5 | 5 | 5 | 5 |
| VR primer | 5 | 5 | 5 | 5 | 5 | 5 | 5 | 5 |
| dNTPmix | 10 | 10 | 10 | 10 | 10 | 10 | 10 | 10 |
| thermo pol buffer | 10 | 10 | 10 | 10 | 10 | 10 | 10 | 10 |
| DNA pol(VENT) | 1 | 1 | 1 | 1 | 1 | 1 | 1 | 1 |
| dH2O | 68 | 68 | 68 | 68 | 68 | 68 | 68 | 68 |
| TOTAL | 100 | 100 | 100 | 100 | 100 | 100 | 100 | 100 |
20,August 2008
TRANSFORMATION
BBa_J52008(Luciferase Renilla)-->colony
BBa_I712019(Luciferase Fire fly)-->no colony
BBa_J332002(LacZ)-->colony
BBa_E2030(Venus YFP)-->not so good
YFP(P-GEX)-->colony
BBa_F2620-->colony
BBa_T9002-->colony
21,August 2008
TRANSFORMATION
BBa_J32007(Lux CDABE)-->no colony
BBa_B0034(RBS)-->colony
22,August 2008
MINI PREP
BBa_J52008(Luciferase)
BBa_F2620
BBa_T9002
BBa_J33202(LacZ)
PUC19
PGEX VENUS YFP(GST fusion)
TRANSFORMATION
PGEX VENUS YFP(GST fusion)
24,August 2008
TRANSFORMATION
BBa_J63001(yeast optimized YFP)-->colony
Digestion
BBa_F2620(1)①
BBa_F2620(2)②
Menbren Venus YFP③
| Sample No | ① | ② | ③ |
| DNA tamplate | 3 | 3 | 3 |
| Buffer3 | 1 | 1 | 1 |
| dH2O | 6 | 6 | 6 |
| EcoRⅠ | 0.5 | 0.5 | 0.5 |
| TOTAL | 10 | 10 | 10 |
->37℃ 30min
| Sample No | ①(afetr digestion) | ②(after digestion) | ③(afetr digestion) |
| DNA tamplate | 10 | 10 | 10 |
| loading Dye | 2 | 2 | 2 |
| TOTAL | 12 | 12 | 12 |
| Sample No | ① | ② | ③ |
| DNA tamplate | 2 | 2 | 2 |
| loading Dye | 1 | 1 | 1 |
| dH2O | 3 | 3 | 3 |
| TOTAL | 6 | 6 | 6 |
->135V 30min
25,August,2008
PCR
BBa_E2030①
BBa_J33202②
BL21③
BBa_J52008④
BBa_I712019⑤
| Sample No | ① | ② | ③ | ④ | ⑤ |
| DNA tamplate | 1 | 1 | 1 | 1 | >1 |
| FW primer | 5(Venus_YFP_fwd) | 5(LacZ_fwd) | 5(LacZ_fwd) | 5(rLUC_fwd) | 5(fLuc_fwd) |
| VR primer | 5(VR) | 5(LacZ_rev) | 5(LacZ_rev) | 5(VR) | 5(VR) |
| dNTPmix | 10 | 10 | 10 | 10 | 10 |
| thermo pol buffer | 10 | 10 | 10 | 10 | 10 |
| DNA pol(VENT) | 1 | 1 | 1 | 1 | 1 |
| dH2O | 68 | 68 | 68 | 68 | 68 |
| TOTAL | 100 | 100 | 100 | 100 | 100 |
->Gel
| Sample No | ① | ② | ③ | ④ | ⑤ |
| DNA tamplate | 1 | 1 | 1 | 1 | >1 |
| loading Dye | 1 | 1 | 1 | 1 | 1 |
| dH2O | 4 | 4 | 4 | 4 | 4 |
| TOTAL | 6 | 6 | 6 | 6 | 6 |
25,August,2008
MINI prep
Venus
BBa_F2620
TRANSFORMATRION
Venus
PUC19
pGFPUV
pGEX Venus YFP
Membren Venus YFP
PCR
BBa_J63001①
| Sample No | ① |
| DNA tamplate | 1 |
| FW primer(Venus) | 5 |
| VR primer | 5 |
| dNTPmix | 10 |
| thermo pol buffer | 10 |
| DNA pol(VENT) | 1 |
| dH2O | 68 |
| TOTAL | 100 |
Digestion
Luciferase(J52008:2007,2008 I712019:2008),Lac Z(J33202:2007,2008),Venus(J32007:2007,2008),Craig Venus,T9002,F2620
- 結果:成功→Luciferase(J52008:2007),Lac Z(J33202:2007),Craig Venus,T9002,F2620
- 失敗→Luciferase(J52008:2008 J52008:2008),Lac Z(J33202:2008),Venus(J32007:2007,2008)
- 2008,8,21(thu)
Lux CDABE(J32007:2007),RBS(B0034:2007)
- 結果:成功→RBS
- 失敗→Lux CDABE(二回行ったがどちらもコロニーができなかった)→三回目に成功
- 2008,8,22(sun)
Venus YFP
- 結果
- 2008,8,22(fri)
J52008(2007),F2620,T9002,PUC19(LacZ),VenusYFP,J33202(2007)
- 結果:ゲル電→J33202はバンドの位置が違っていましたが、大丈夫だと思われます。他は正しいバンドがでました。
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