Team:Chiba/jk/γ

From 2008.igem.org

(Difference between revisions)

Revision as of 11:02, 16 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

</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	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_J52008①
vector:BBa_F2620 insert:BBa_E2030②
Negative control:BBa_F2620③

Sample No	①	②	③
vector	5	5	5
inser	10	7.5	0
Ligase Buffer	4	4	4
Ligase	1	1	1
dH2O	0	2.5	10
TOTAL	20	20	20

->RT 2hour
->TRANSFORMATION XL10G
->37℃ over night
->colony PCR
->MINI PREP

28,August,2008
TIME RESPONCE(liquid)

29,August,2008
TIME RESPONCE(solid)

30,August,2008
TRANSFORMATION
mcherry
PCR
BBa_J52008(rLuc)

</tr>

Sample No	BBa_J52008
DNA tamplate	1
rLuc_fwd	2.5
VR	2.5
thermo pol Buffer	5
dNTP	5
Vent pol	0.5
dH2O	34
TOTAL	50

31,August,2008
digestion
BBa_R0010①
PCR product(BBa_J52008)②

Sample No	①	②
DNA	10	10
SpeⅠ	1	0
PstⅠ	1	1
XbaⅠ	0	1
BSA(x100)	0.5	0.5
Buffer2	5	0
Buffer3	0	5
dH20	28.5	28.5
TOTAL	50	50

->37℃ 3hour
->①20μl refine
->②20μl refine

1,September,2008
Ligation
vector:BBa_R0010 insert:BBa_J52008①
negative control:BBa_R0010②

</tr>

Sample No	①	②
vector	2	2
insert	3	0
Ligase Buffer	2	2
Ligase	1	1
dH2O	3	5
TOTAL	10	10

->R/T 2hour

TRANSFORMATION
BBa_R0079(Las promoter)
BBa_R0071(RhlR promoter)
BBa_R0077(cinR promoter+RBS)
BBa_R0078(cnR promoter)
BBa_R0062(LUX promoter)

2,September,2008
COLONY PCR
BBa_R0010+BBa_J52008①②③④⑤⑥⑦⑧⑨⑩⑪⑫⑬⑭⑮

</tr>

Sample No

①

②

③

④

⑤

⑥

⑦

⑧

⑨

⑩

⑪

⑫

⑬

⑭

⑮

①

②

③

④

⑤

⑥

⑦

⑧

⑨

⑩

⑪

⑫

⑬

⑭

⑮

SpeⅠ

2

PstⅠ

2

BSA(x10)

13

Buffer2

13

TOTAL

130

ホーム	メンバー紹介	プロジェクト紹介	Parts Submitted to the Registry	モデリング	ノート

@@ Line 1: / Line 1: @@
+----
 '''17,August 2008'''
 <br>PCR
@@ Line 48: / Line 49: @@
 </table>
+----
 '''20,August 2008'''
 <br>TRANSFORMATION
@@ Line 59: / Line 62: @@
+----
 '''21,August 2008'''
 <br>TRANSFORMATION
@@ Line 64: / Line 68: @@
 <br>BBa_B0034(RBS)-->colony
+----
 '''22,August 2008'''
 <br>MINI PREP
@@ Line 76: / Line 82: @@
 <br>PGEX VENUS YFP(GST fusion)
+----
 '''24,August 2008'''
 <br>TRANSFORMATION
@@ Line 152: / Line 159: @@
 <br>->135V 30min
+----
 '''25,August,2008'''
 <br>PCR
@@ Line 270: / Line 278: @@
 </tr>
 </table>
+----
 '''26,August,2008'''
 <br>Gel check
@@ Line 408: / Line 418: @@
 <br>->MINI PREP
+----
 '''28,August,2008'''
 <br>TIME RESPONCE(liquid)
+----
 '''29,August,2008'''
 <br>TIME RESPONCE(solid)
+----
 '''30,August,2008'''
 <br>TRANSFORMATION
@@ Line 457: / Line 470: @@
 </table>
+----
 '''31,August,2008'''
 <br>digestion
@@ Line 481: / Line 495: @@
 <td>XbaⅠ</td>
 <td>0</td><td>1</td>
+</tr>
+<tr>
 <td>BSA(x100)</td>
 <td>0.5</td><td>0.5</td>
@@ Line 495: / Line 511: @@
 <td>dH20</td>
 <td>28.5</td><td>28.5</td>
+</tr>
+<tr>
 <td>TOTAL</td>
 <td>50</td><td>50</td>
@@ Line 503: / Line 521: @@
 <br>->②20μl refine
+----
 '''1,September,2008'''
 <br>Ligation
@@ Line 538: / Line 557: @@
 <br>->R/T 2hour
+<br>TRANSFORMATION
+<br>BBa_R0079(Las promoter)
+<br>BBa_R0071(RhlR promoter)
+<br>BBa_R0077(cinR promoter+RBS)
+<br>BBa_R0078(cnR promoter)
+<br>BBa_R0062(LUX promoter)
+----
+'''2,September,2008'''
+<br>COLONY PCR
+<br>BBa_R0010+BBa_J52008①②③④⑤⑥⑦⑧⑨⑩⑪⑫⑬⑭⑮
+<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><td>⑨</td><td>⑩</td><td>⑪</td><td>⑫</td><td>⑬</td><td>⑭</td><td>⑮</td>
+</tr>
+<tr>
+<td></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><td>⑨</td><td>⑩</td><td>⑪</td><td>⑫</td><td>⑬</td><td>⑭</td><td>⑮</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>