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Team:Chiba/jk/γ
From 2008.igem.org
| Line 278: | Line 278: | ||
<tr> | <tr> | ||
<td width="257">Sample No</td> | <td width="257">Sample No</td> | ||
| - | <td>①</td><td>②</td><td>③</td><td>④</td><td>⑤ | + | <td>①</td><td>②</td><td>③</td><td>④</td><td>⑤</td> |
</tr> | </tr> | ||
<tr> | <tr> | ||
| Line 290: | Line 290: | ||
<tr> | <tr> | ||
<td>dH2O</td> | <td>dH2O</td> | ||
| - | <td>4</td><td>4</td><td>4</td><td>4</td><td></td> | + | <td>4</td><td>4</td><td>4</td><td>4</td><td>4</td> |
</tr> | </tr> | ||
<tr> | <tr> | ||
| Line 370: | Line 370: | ||
<br>Ligation | <br>Ligation | ||
<br>vector:BBa_F2620 insert:BBa_J52008 | <br>vector:BBa_F2620 insert:BBa_J52008 | ||
| + | <table width="250" border="4" cellpadding="0" cellspacing="0" bordercolor="#000000"> | ||
| + | <tr> | ||
| + | <td width="257">Sample No</td> | ||
| + | <td>①</td> | ||
| + | </tr> | ||
| + | <tr> | ||
| + | <td>insert</td> | ||
| + | <td>1</td> | ||
| + | </tr> | ||
| + | <tr> | ||
| + | <td>vector</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></td> | ||
| + | </tr> | ||
| + | <tr> | ||
| + | <td>TOTAL</td> | ||
| + | <td>6</td><td>6</td><td>6</td><td>6</td><td>6</td> | ||
| + | </tr> | ||
| + | </table> | ||
| + | |||
| + | |||
Revision as of 10:37, 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 |
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 |
26,August,2008
Gel check
BBa_F2620
BBa_F2620
BBa_J63001
| 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 |
Digestion
vector:BBa_F2620
| Sample No | BBa_F2620 |
| DNA tamplate | 100 |
| SpeⅠ | 2 |
| PstⅠ | 2 |
| BSA(x10) | 13 |
| Buffer2 | 13 | TOTAL | 130 |
insert:BBa_J52008,BBa_E2030
| Sample No | J52008 | E2030 |
| DNA tamplate | 30 | 30 |
| XbaⅠ | 1 | 1 |
| PstⅠ | 1 | 1 |
| DpnⅠ | 1 | 1 |
| Buffer3 | 4 | 4 |
| BSA(x10) | 4 | 4 | TOTAL | 40 | 40 |
->37℃ 3hour
->SAP
->37℃ 30min
->65℃ 15min
->Zymo Clean
Ligation
vector:BBa_F2620 insert:BBa_J52008
| Sample No | ① | ||||
| insert | 1 | ||||
| vector | 1 | 1 | 1 | 1 | 1 |
| dH2O | 4 | 4 | 4 | 4 | |
| TOTAL | 6 | 6 | 6 | 6 | 6 |
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