Team:Chiba/jk/γ
From 2008.igem.org
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'''22,August 2008''' | '''22,August 2008''' | ||
<br>MINI PREP | <br>MINI PREP | ||
- | <br>BBa_J52008 | + | <br>BBa_J52008(Luciferase) |
<br>BBa_F2620 | <br>BBa_F2620 | ||
<br>BBa_T9002 | <br>BBa_T9002 | ||
+ | <br>BBa_J33202(LacZ) | ||
+ | <br>PUC19 | ||
+ | <br>PGEX VENUS YFP(GST fusion) | ||
<br> | <br> | ||
+ | <br>TRANSFORMATION | ||
+ | <br>PGEX VENUS YFP(GST fusion) | ||
+ | |||
'''24,August 2008''' | '''24,August 2008''' | ||
<br>TRANSFORMATION | <br>TRANSFORMATION | ||
<br>BBa_J63001(yeast optimized YFP)-->colony | <br>BBa_J63001(yeast optimized YFP)-->colony | ||
- | + | <br>Digestion | |
+ | <br>BBa_F2620(1)① | ||
+ | <br>BBa_F2620(2)② | ||
+ | <br>Menbren Venus YFP③ | ||
+ | <table width="250" border="4" cellpadding="0" cellspacing="0" bordercolor="#000000"> | ||
+ | <tr> | ||
+ | <td width="257">Sample No</td> | ||
+ | <td>①</td><td>②</td><td>③</td> | ||
+ | </tr> | ||
+ | <tr> | ||
+ | <td>DNA tamplate</td> | ||
+ | <td>3</td><td>3</td><td>3</td> | ||
+ | </tr> | ||
+ | <tr> | ||
+ | <td>Buffer3</td> | ||
+ | <td>1</td><td>1</td><td>1</td> | ||
+ | </tr> | ||
+ | <tr> | ||
+ | <td>dH2O</td> | ||
+ | <td>6</td><td>6</td><td>6</td> | ||
+ | </tr> | ||
+ | <tr> | ||
+ | <td>EcoRⅠ</td> | ||
+ | <td>0.5</td><td>0.5</td><td>0.5</td> | ||
+ | </tr> | ||
+ | <tr> | ||
+ | <td>TOTAL</td> | ||
+ | <td>10</td><td>10</td><td>10</td> | ||
+ | </tr> | ||
+ | </table> | ||
+ | <br>->37℃ 30min | ||
+ | <table width="250" border="4" cellpadding="0" cellspacing="0" bordercolor="#000000"> | ||
+ | <tr> | ||
+ | <td width="257">Sample No</td> | ||
+ | <td>①(afetr digestion)</td><td>②(after digestion)</td><td>③(afetr digestion)</td> | ||
+ | </tr> | ||
+ | <tr> | ||
+ | <td>DNA tamplate</td> | ||
+ | <td>10</td><td>10</td><td>10</td> | ||
+ | </tr> | ||
+ | <tr> | ||
+ | <td>loading Dye</td> | ||
+ | <td>2</td><td>2</td><td>2</td> | ||
+ | </tr> | ||
+ | <tr> | ||
+ | <td>TOTAL</td> | ||
+ | <td>12</td><td>12</td><td>12</td> | ||
+ | </tr> | ||
+ | </table> | ||
+ | <table width="250" border="4" cellpadding="0" cellspacing="0" bordercolor="#000000"> | ||
+ | <tr> | ||
+ | <td width="257">Sample No</td> | ||
+ | <td>①</td><td>②</td><td>③</td> | ||
+ | </tr> | ||
+ | <tr> | ||
+ | <td>DNA tamplate</td> | ||
+ | <td>2</td><td>2</td><td>2</td> | ||
+ | </tr> | ||
+ | <tr> | ||
+ | <td>loading Dye</td> | ||
+ | <td>1</td><td>1</td><td>1</td> | ||
+ | </tr> | ||
+ | <tr> | ||
+ | <td>dH2O</td> | ||
+ | <td>3</td><td>3</td><td>3</td> | ||
+ | </tr> | ||
+ | <tr> | ||
+ | <td>TOTAL</td> | ||
+ | <td>6</td><td>6</td><td>6</td> | ||
+ | </tr> | ||
+ | </table> | ||
+ | <br>->135V 30min | ||
+ | |||
+ | '''25,August,2008''' | ||
+ | <br>PCR | ||
+ | <br>BBa_E2030① | ||
+ | <br>BBa_J33202② | ||
+ | <br>BL21③ | ||
+ | <br>BBa_J52008④ | ||
+ | <br>BBa_I712019⑤ | ||
+ | <table width="250" border="4" cellpadding="0" cellspacing="0" bordercolor="#000000"> | ||
+ | <tr> | ||
+ | <td width="257">Sample No</td> | ||
+ | <td>①</td><td>②</td><td>③</td><td>④</td><td>⑤</td> | ||
+ | </tr> | ||
+ | <tr> | ||
+ | <td>DNA tamplate</td> | ||
+ | <td>1</td><td>1</td><td>1</td><td>1</td>><td>1</td> | ||
+ | </tr> | ||
+ | <tr> | ||
+ | <td>FW primer</td> | ||
+ | <td>5(Venus_YFP_fwd)</td><td>5(LacZ_fwd)</td><td>5(LacZ_fwd)</td><td>5(rLUC_fwd)</td><td>5(fLuc_fwd)</td> | ||
+ | </tr> | ||
+ | <tr> | ||
+ | <td>VR primer</td> | ||
+ | <td>5(VR)</td><td>5(LacZ_rev)</td><td>5(LacZ_rev)</td><td>5(VR)</td><td>5(VR)</td> | ||
+ | </tr> | ||
+ | <tr> | ||
+ | <td>dNTPmix</td> | ||
+ | <td>10</td><td>10</td><td>10</td><td>10</td><td>10</td> | ||
+ | </tr> | ||
+ | <tr> | ||
+ | <td>thermo pol buffer</td> | ||
+ | <td>10</td><td>10</td><td>10</td><td>10</td><td>10</td> | ||
+ | </tr> | ||
+ | <tr> | ||
+ | <td>DNA pol(VENT)</td> | ||
+ | <td>1</td><td>1</td><td>1</td><td>1</td><td>1</td> | ||
+ | </tr> | ||
+ | <tr> | ||
+ | <td>dH2O</td> | ||
+ | <td>68</td><td>68</td><td>68</td><td>68</td><td>68</td> | ||
+ | </tr> | ||
+ | <tr> | ||
+ | <td>TOTAL</td> | ||
+ | <td>100</td><td>100</td><td>100</td><td>100</td><td>100</td> | ||
+ | </tr> | ||
+ | </table> | ||
+ | <br>->Gel | ||
+ | <table width="250" border="4" cellpadding="0" cellspacing="0" bordercolor="#000000"> | ||
+ | <tr> | ||
+ | <td width="257">Sample No</td> | ||
+ | <td>①</td><td>②</td><td>③</td><td>④</td><td>⑤</td> | ||
+ | </tr> | ||
+ | <tr> | ||
+ | <td>DNA tamplate</td> | ||
+ | <td>1</td><td>1</td><td>1</td><td>1</td>><td>1</td> | ||
+ | </tr> | ||
+ | <tr> | ||
+ | <td>loading Dye</td> | ||
+ | <td>1</td><td>1</td><td>1</td><td>1</td><td>1</td> | ||
+ | </tr> | ||
+ | <tr> | ||
+ | <td>dH2O</td> | ||
+ | <td>4</td><td>4</td><td>4</td><td>4</td><td>4</td> | ||
+ | </tr> | ||
+ | <tr> | ||
+ | <td>TOTAL</td> | ||
+ | <td>6</td><td>6</td><td>6</td><td>6</td><td>6</td> | ||
+ | </tr> | ||
+ | </table> | ||
+ | |||
+ | '''25,August,2008''' | ||
+ | <br>MINI prep | ||
+ | <br>Venus | ||
+ | <br>BBa_F2620 | ||
+ | <br>TRANSFORMATRION | ||
+ | <br>Venus | ||
+ | <br>PUC19 | ||
+ | <br>pGFPUV | ||
+ | <br>pGEX Venus YFP | ||
+ | <br>Membren Venus YFP | ||
+ | <br>PCR | ||
+ | <br>BBa_J63001① | ||
+ | <table width="250" border="4" cellpadding="0" cellspacing="0" bordercolor="#000000"> | ||
+ | <tr> | ||
+ | <td width="257">Sample No</td> | ||
+ | <td>①</td> | ||
+ | </tr> | ||
+ | <tr> | ||
+ | <td>DNA tamplate</td> | ||
+ | <td>1</td> | ||
+ | </tr> | ||
+ | <tr> | ||
+ | <td>FW primer(Venus)</td> | ||
+ | <td>5</td> | ||
+ | </tr> | ||
+ | <tr> | ||
+ | <td>VR primer</td> | ||
+ | <td>5</td> | ||
+ | </tr> | ||
+ | <tr> | ||
+ | <td>dNTPmix</td> | ||
+ | <td>10</td> | ||
+ | </tr> | ||
+ | <tr> | ||
+ | <td>thermo pol buffer</td> | ||
+ | <td>10</td> | ||
+ | </tr> | ||
+ | <tr> | ||
+ | <td>DNA pol(VENT)</td> | ||
+ | <td>1</td> | ||
+ | </tr> | ||
+ | <tr> | ||
+ | <td>dH2O</td> | ||
+ | <td>68</td> | ||
+ | </tr> | ||
+ | <tr> | ||
+ | <td>TOTAL</td> | ||
+ | <td>100</td> | ||
+ | </tr> | ||
+ | </table> | ||
+ | <br>Digestion | ||
+ | |||
+ | |||
+ | |||
+ | |||
Luciferase(J52008:2007,2008 I712019:2008),Lac Z(J33202:2007,2008),Venus(J32007:2007,2008),Craig Venus,T9002,F2620 | 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 | :結果:成功→Luciferase(J52008:2007),Lac Z(J33202:2007),Craig Venus,T9002,F2620 | ||
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Venus YFP | Venus YFP | ||
:結果 | :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。 | ||
- | |||
- | |||
- | |||
- | |||
{| style="color:white;background-color:Maroon" cellpadding="3" cellspacing="3" border="1" bordercolor="white" width="100%" align="center" | {| style="color:white;background-color:Maroon" cellpadding="3" cellspacing="3" border="1" bordercolor="white" width="100%" align="center" | ||
!align="center"|[[Team:Chiba/j|ホーム]] | !align="center"|[[Team:Chiba/j|ホーム]] |
Revision as of 07:45, 14 September 2008
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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