Team:Chiba/jk/γ

From 2008.igem.org

(Difference between revisions)
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</table>
</table>
-
'''25,August,2008'''
 
<br>MINI prep
<br>MINI prep
<br>Venus
<br>Venus
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</tr>
</tr>
</table>
</table>
 +
'''26,August,2008'''
 +
<br>Gel check
 +
<br>BBa_F2620
 +
<br>BBa_F2620
 +
<br>BBa_J63001
 +
<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><td>⑥</td><td>⑦</td><td>⑧</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></td>
 +
</tr>
 +
<tr>
 +
<td>TOTAL</td>
 +
<td>6</td><td>6</td><td>6</td><td>6</td><td>6</td>
 +
</tr>
 +
</table>
 +
<br>Digestion
<br>Digestion
 +
<br>vector:BBa_F2620
 +
<table width="250" border="4" cellpadding="0" cellspacing="0" bordercolor="#000000">
 +
<tr>
 +
<td width="257">Sample No</td>
 +
<td>BBa_F2620</td>
 +
</tr>
 +
<tr>
 +
<td>DNA tamplate</td>
 +
<td>100</td>
 +
</tr>
 +
<tr>
 +
<td>SpeⅠ</td>
 +
<td>2</td>
 +
</tr>
 +
<tr>
 +
<td>PstⅠ</td>
 +
<td>2</td>
 +
</tr>
 +
<tr>
 +
<td>BSA(x10)</td>
 +
<td>13</td>
 +
</tr>
 +
<tr>
 +
<td>Buffer2</td>
 +
<td>13</td>
 +
</tr>
 +
<td>TOTAL</td>
 +
<td>130</td>
 +
</tr>
 +
</table>
 +
<br>insert:BBa_J52008,BBa_E2030
 +
<table width="250" border="4" cellpadding="0" cellspacing="0" bordercolor="#000000">
 +
<tr>
 +
<td width="257">Sample No</td>
 +
<td>J52008</td><td>E2030</td>
 +
</tr>
 +
<tr>
 +
<td>DNA tamplate</td>
 +
<td>30</td><td>30</td>
 +
</tr>
 +
<tr>
 +
<td>XbaⅠ</td>
 +
<td>1</td><td>1</td>
 +
</tr>
 +
<tr>
 +
<td>PstⅠ</td>
 +
<td>1</td><td>1</td>
 +
</tr>
 +
<tr>
 +
<td>DpnⅠ</td>
 +
<td>1</td><td>1</td>
 +
</tr>
 +
<tr>
 +
<td>Buffer3</td>
 +
<td>4</td><td>4</td>
 +
</tr>
 +
<tr>
 +
<td>BSA(x10)</td>
 +
<td>4</td><td>4</td>
 +
</tr>
 +
<td>TOTAL</td>
 +
<td>40</td><td>40</td>
 +
</tr>
 +
</table>
 +
<br>->37℃　3hour
 +
<br>->SAP
 +
<br>->37℃ 30min
 +
<br>->65℃ 15min
 +
<br>->Zymo Clean
 +
<br>Ligation
 +
<br>vector:BBa_F2620 insert:BBa_J52008
-
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。
 

Revision as of 10:04, 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 ①②③④⑤⑥⑦⑧
DNA tamplate 11111111
FW primer 55555555
VR primer 55555555
dNTPmix 1010101010101010
thermo pol buffer 1010101010101010
DNA pol(VENT) 11111111
dH2O 6868686868686868
TOTAL 100100100100100100100100

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 ①②③
DNA tamplate 333
Buffer3 111
dH2O 666
EcoRⅠ 0.50.50.5
TOTAL 101010


->37℃ 30min

Sample No ①(afetr digestion)②(after digestion)③(afetr digestion)
DNA tamplate 101010
loading Dye 222
TOTAL 121212
Sample No ①②③
DNA tamplate 222
loading Dye 111
dH2O 333
TOTAL 666


->135V 30min

25,August,2008
PCR
BBa_E2030①
BBa_J33202②
BL21③
BBa_J52008④
BBa_I712019⑤

>
Sample No ①②③④⑤
DNA tamplate 11111
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 1010101010
thermo pol buffer 1010101010
DNA pol(VENT) 11111
dH2O 6868686868
TOTAL 100100100100100


->Gel

>
Sample No ①②③④⑤
DNA tamplate 11111
loading Dye 11111
dH2O 44444
TOTAL 66666


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 ①②③④⑤⑥⑦⑧
DNA tamplate 11111
Loading Dye 11111
dH2O 4444
TOTAL 66666


Digestion
vector:BBa_F2620

</tr>

Sample No BBa_F2620
DNA tamplate 100
SpeⅠ 2
PstⅠ 2
BSA(x10) 13
Buffer2 13
TOTAL 130


insert:BBa_J52008,BBa_E2030

</tr>

Sample No J52008E2030
DNA tamplate 3030
XbaⅠ 11
PstⅠ 11
DpnⅠ 11
Buffer3 44
BSA(x10) 44
TOTAL 4040


->37℃　3hour
->SAP
->37℃ 30min
->65℃ 15min
->Zymo Clean
Ligation
vector:BBa_F2620 insert:BBa_J52008





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