Template:Kyoto/Notebook/Sep 26

From 2013.igem.org

(Difference between revisions)

Revision as of 07:13, 27 September 2013

Sep 26

Electrophoresis

No name

Lane	Sample	Enzyme1	Enzyme2
1	GGA1-1
2	GGA1-2
3	GGA1-3
4	GGA1-4
5	GGA1-5
6	GGA1-6
7	GGA1-7
8	GGA1-8
9	1kbp ladder
10	GGA2-1
11	GGA2-2
12	GGA2-3
13	GGA2-4
14	GGA2-5
15	GGA2-6
16	GGA2-7
17	GGA2-8

Lane	Sample	Enzyme1	Enzyme2
1	GGA3-1
2	GGA3-2
3	GGA3-3
4	1kbp ladder
5	GGA3-4
6	GGA3-5
7	GGA3-6
8	GGA3-7

</div>

Lane	Sample	Enzyme1	Enzyme2
1	GGA3-8
2	GGA4-1
3	GGA4-2
4	GGA4-3
5	1kbp ladder
6	GGA4-4
7	GGA4-5
8	GGA4-6

</div>

Lane	Sample	Enzyme1	Enzyme2
1	GGA4-7
2	GGA4-8
3	1kbp ladder
4	GGA7-1
5	GGA16-1
6	NC(RBS-GFP-DT)

</div>

Ligasion(Golden Gate Assenbly)

No name

state	Vector		Inserter1		Inserter2		Inserter3		Inserter4		Inserter5		(NEB)T4 ligase	(NEB)10*T4 ligase	MilliQ
experiment	RBS-GFP-DT	1.0 µL	Pcon-pT181 antisense(E-2A)	0.6µL	DT(2-1)	0.5µL	Pcon-pT181 attenuator(1-SA)	2.1 µL	--	--	--	--	0.5 µL	0.5 µL	up to 5 µL
experiment	pSB1C3(EcoRI&SpeI)	1.8 µL	Pcon-pT181 antisense(E-2A)	0.9 µL	DT(2-1)	0.5 µL	Pcon-GFP-DT(1-SA)	1.5 µL	--	--	--	--	0.5 µL	0.5 µL	up to 5 µL
experiment	RBS-GFP-DT	1.0 µL	Ptet(E-0)	0.7µL	Pcon pT181 attenuator(0-1A)	0.3µL	tet aptamer(1-3)	0.3 µL	Pcon-tetR-DT(3-2A)	1.1 µL	Ptet(2-5)	0.3 µL	0.5 µL	0.5 µL	up to 5 µL
experiment	RBS-GFP-DT	1.0 µL	Ptet(E-0)	0.7 µL	Pcon-pT181 attenuator-DT(0-1A)	1.0 µL	Pcon-tetR-DT(1-2A)	1.0 µL	Ptet(2-5)	0.3µL	--	--	0.5 µL	0.5 µL	up to 5 µL
experiment	RBS-GFP-DT	1.0 µL	Pcon-pT181 attenuator-tetR-DT(E-1A)	1.0 µL	Pcon antisense-spinach-DT(1-2A)	0.7µL	Pcon-pT 181 attenuator(2-SA)	2.1 µL	--	--	--	--	0.5 µL	0.5 µL	up to 5 µL
experiment	RBS-GFP-DT	1.0 µL	Pcon antisense(E-1A)	0.5 µL	spinach-DT(1-2)	1.0 µL	Pcon-GFP-DT(1-2)	0.8 µL	--	--	--	--	0.5 µL	0.5 µL	up to 5 µL
experiment	pSB1C3(EcoRI&SpeI)	1.8 µL	Pcon-pT181 antisense(E-1A)	0.8µL	spinach-DT(1-2)	1.0µL	Pcon-GFP-DT(2-SA)	0.8 µL	--	--	--	--	0.5 µL	0.5 µL	up to 5 µL
experiment	pSB1C3(EcoRI&SpeI)	1.8 µL	Pcon-tet aptamer-DT(E-1A)	3.0 µL	Ptet-pT181 antisense(1-3)	1.5 µL	spinach-DT(3-2)	1.5 µL	Pcon-GFP-DT(2-5A)	0.8 µL	--	--	0.5 µL	0.5 µL	up to 5 µL

Liquid Culture

No name

Sample	medium
Pcon-GFP-DT	Amp
RBS-GFP-DT	CP
Pcon-pT181 attenuator-DT	Amp
Pcon-tet aptamer-DT	Amp
Pcon-spinach-DT	Amp
Pcon-pT181 antisense-spinach-DT	Amp
spinach-DT	CP

BSAI Digestion

No name

	DNA	MilliQ	BSAI-HF	10*cut smart	total
Pcon	7 µL	10.7 µL	0.3 µL	2 µL	20 µL
RBS-luxR-DT	10 µL	7 µL	0.3 µL	2 µL	20 µL

Colony PCR

Tatsui

Sample	base pair
GGA-1 (9~16)	3177
GGA-2 (9~16)	3189
GGA-3 (9~21)	2989
GGA-4 (9~16)	2939
GGA-6 (1)	3359
GGA-16(2~4)	2939

PreDenature	Denature	Annealing	Extension	cycle
94°C	94°C	55°C	68°C	--
5min	30s	30s	3min30s	30cycles

Electrophoresis

No name

Lane	Sample	Enzyme1	Enzyme2
1	GGA-1 (9)	--	--
2	GGA-1 (10)	--	--
3	GGA-1 (11)	--	--
4	GGA-1 (12)	--	--
5	GGA-1 (13)	--	--
6	GGA-1 (14)	--	--
7	GGA-1 (15)	--	--
8	GGA-1 (16)	--	--
9	1kbp ladder	--	--
10	GGA-2 (9)	--	--
11	GGA-2 (10)	--	--
12	GGA-2 (11)	--	--
13	GGA-2 (12)	--	--
14	GGA-2 (13)	--	--
15	GGA-2 (14)	--	--
16	GGA-2 (15)	--	--
17	GGA-2 (16)	--	--

Lane	Sample	Enzyme1	Enzyme2
1	GGA-4 (9)	--	--
2	GGA-4 (10)	--	--
3	GGA-4 (11)	--	--
4	GGA-4 (12)	--	--
5	GGA-4 (13)	--	--
6	GGA-4 (14)	--	--
7	GGA-4 (15)	--	--
8	GGA-4 (16)	--	--
9	1kbp ladder	--	--
10	GGA-6 (1)	--	--
11	GGA-16 (2)	--	--
12	GGA-16 (3)	--	--
13	GGA-16 (4)	--	--

</div>

Lane	Sample	Enzyme1	Enzyme2
1	GGA-3 (9)	--	--
2	GGA-3 (10)	--	--
3	GGA-3 (11)	--	--
4	GGA-3 (12)	--	--
5	1kbp ladder	--	--
6	GGA-3 (13)	--	--
7	GGA-3 (15)	--	--
8	GGA-3 (14)	--	--

</div>

Lane	Sample	Enzyme1	Enzyme2
1	GGA-3 (17)	--	--
2	GGA-3 (18)	--	--
3	GGA-3 (19)	--	--
4	1kbp ladder	--	--
5	GGA-3 (20)	--	--
6	GGA-3 (21)	--	--
7	GGA-3 (16)	--	--

</div>

Electrophoresis

No name

Lane	Sample	Enzyme1	Enzyme2
1	RBS-GFP-DT(RNA)	--	--
2	1kbp ladder	--	--
3	Pcon-RBS-GFP-DT(RNA)	--	--

Liquid Culture

Hirano

Sample	medium
Pcon-GFP-DT	--
RBS-GFP-DT	--
Pcon-tet aptamer-DT	--
Pcon-spinach-DT	--
Pcon-pT181 antisense-spinach-DT	--
spinach-DT	--

Colony PCR

Hirano

Sample	base pair
9/25 Pcon-pH181 attenuator+RBS-GFP-DT-1	--
9/25 Pcon-pH181 attenuator+RBS-GFP-DT-2	--
9/25 Pcon-pH181 attenuator+RBS-GFP-DT-3	--
9/25 Pcon-pH181 attenuator+RBS-GFP-DT-4	--
9/24 Ptet-RBS-GFP-DT-1	--

PreDenature	Denature	Annealing	Extension	cycle
94°C	94°C	55°C	68°C	--
5min	30s	30s	1min40s	30cycles

Ligasion(Golden Gate Assenbly)

Hirano

state	Vector	Inserter1	Inserter2	Inserter3	Inserter4
experiment	GGA2	Pcon-attenuator(E-1A)	Pcon-tetR-DT(1-2A)
experiment	GGA3	Pcon-tetap-DT(E-1A)	Pcon-tetR-DT(1-2A)	Pcon-GFP-DT(1-SA)	1.5 µL	--	--	--	--	0.5 µL	0.5 µL	up to 5 µL
experiment	GGA4	Pcon-spi-DT(E-1A)	Pcon-tetR-DT(1-2A)	tet aptamer(1-3)	0.3 µL	Pcon-tetR-DT(3-2A)	1.1 µL
experiment	GGA5	Pcon-antisense(E-1A)	Pcon-pT181 attenuator-DT(0-1A)	1.0 µL	Pcon-tetR-DT(1-2A)	1.0 µL	Ptet(2-5)	0.3µL	--	--	0.5 µL	0.5 µL	up to 5 µL
experiment	GGA6	Pcon-attenuator(E-1A)	1.0 µL	Pcon antisense-spinach-DT(1-2A)	0.7µL	Pcon-pT 181 attenuator(2-SA)	2.1 µL	--	--	--	--	0.5 µL	0.5 µL	up to 5 µL
experiment	GGA7	Pcon-tetR-DT(E-2A)	0.5 µL	spinach-DT(1-2)	1.0 µL	Pcon-GFP-DT(1-2)	0.8 µL	--	--	--	--	0.5 µL	0.5 µL	up to 5 µL

Plating

Hirano

Sample	Use plate
9/26 GGA2	LB(CP)
9/26 GGA3	LB(CP)
9/26 GGA4	LB(CP)
9/26 GGA5	LB(CP)
9/26 GGA6	LB(CP)
9/26 GGA7	LB(CP)
9/26 GGA11	LB(CP)
9/26 GGA312	LB(CP)
9/26 GGA13	LB(CP)
9/26 GGA14	LB(CP)
9/26 GGA15	LB(CP)
9/26 GGA20	LB(CP)
9/26 GGA23	LB(CP)
9/26 GGA24	LB(CP)
9/26 Ptet+RBS-GFP-DT	LB(CP)

</div> incubate 37°C

Restriction Enzyme Digestion

Kojima

	9/24 pSB1C3	EcoRI	SpeI	buffer	MilliQ	total
2cuts	11µL	1µL	1µL	3µL	14µL	30µL
NC	0.5µL	0µL	0µL	1µL	8.5µL	10µL

	9/24 Pcon-pT181attenuator-aptamer12-1R-DT	EcoRI	SpeI	buffer	MilliQ	total
2cuts	8.7µL	1µL	1µL	3µL	16.3µL	30µL
NC	0.4µL	0µL	0µL	1µL	8.6µL	10µL

	9/24 pT181attenuator	EcoRI	SpeI	buffer	MilliQ	total
2cuts	3.1µL	1µL	1µL	3µL	21.9µL	30µL
NC	0.2µL	0µL	0µL	1µL	8.8µL	10µL

	8/17 RBS-GFP-DT	EcoRI	BSA	buffer	MilliQ	total
1cut	16.4µL	1µL	3µL	3µL	6.6µL	30µL
NC	0.4µL	0µL	1µL	1µL	7.6µL	10µL

	9/17 pSB4K5	EcoRI	PstI	BSA	buffer	MilliQ	total
2cuts	8.5µL	1µL	1µL	3µL	3µL	13.5µL	30µL
NC	0.4µL	0µL	0µL	1µL	1µL	7.6µL	10µL

	9/14 pSB4K5	EcoRI	PstI	BSA	buffer	MilliQ	total
2cuts	7.3µL	1µL	1µL	3µL	3µL	14.7µL	30µL
NC	0.4µL	0µL	0µL	1µL	1µL	7.6µL	10µL

	9/22 Pcon-pT181attenuator-DT	EcoRI	SpeI	buffer	MilliQ	total
2cuts	5.4µL	1µL	1µL	3µL	19.6µL	30µL
NC	0.3µL	0µL	0µL	1µL	8.7µL	10µL

	8/7 RBS-GFP-DT 1	XbaI	PstI	BSA	buffer	MilliQ	total
2cuts	8.1µL	1µL	1µL	3µL	3µL	13.9µL	30µL
NC	0.4µL	0µL	0µL	1µL	1µL	7.6µL	10µL

	9/26 RBS-GFP-DT 1	XbaI	BSA	buffer	MilliQ	total
1cut	8.1µL	1µL	3µL	3µL	14.9µL	30µL

	9/16 Pλ-luxI	EcoRI	PstI	BSA	buffer	MilliQ	total
2cuts	14.7µL	1µL	1µL	3µL	3µL	7.3µL	30µL
NC	0.7µL	0µL	0µL	1µL	1µL	7.3µL	10µL

	9/21 Ptet	EcoRI	SpeI	buffer	MilliQ	total
2cuts	14.2µL	1µL	1µL	3µL	10.8µL	30µL
NC	0.7µL	0µL	0µL	1µL	8.3µL	10µL

	9/24 Pcon-pT181attenuator-aptamer12-1R-DT	XbaI	PstI	BSA	buffer	MilliQ	total
2cuts	8.7µL	1µL	1µL	3µL	3µL	13.3µL	30µL
NC	0.4µL	0µL	0µL	1µL	1µL	7.6µL	10µL

Electrophoresis

Kojima

Lane	Sample	Enzyme1	Enzyme2
1	pSB1C3	EcoRI	SpeI
2	pSB1C3	--	--
3	Pcon-pT181attenuator-aptamer12-1R-DT	EcoRI	SpeI
4	Pcon-pT181attenuator-aptamer12-1R-DT	--	--
5	1kb ladder	--	--
6	pT181attenuator	EcoRI	SpeI
7	pT181attenuator	--	--
8	RBS-GFP-DT	EcoRI	--
9	RBS-GFP-DT	--	--

File:IgkuxxxxBa 1.jpg

@@ Line 385: / Line 385: @@
 |}
 </div>
+===Plating===
+<div class="experiment">
+<span class="author">Hirano</span>
+</div>
+{| class="wikitable"
+!Sample||Use plate
+|-
+|9/26 GGA2 ||LB(CP)
+|-
+|9/26 GGA3 ||LB(CP)
+|-
+|9/26 GGA4||LB(CP)
+|-
+|9/26 GGA5 ||LB(CP)
+|-
+|9/26 GGA6 ||LB(CP)
+|-
+|9/26 GGA7 ||LB(CP)
+|-
+|9/26 GGA11 ||LB(CP)
+|-
+|9/26 GGA312||LB(CP)
+|-
+|9/26 GGA13 ||LB(CP)
+|-
+|9/26 GGA14||LB(CP)
+|-
+|9/26 GGA15 ||LB(CP)
+|-
+|9/26 GGA20||LB(CP)
+|-
+|9/26 GGA23 ||LB(CP)
+|-
+|9/26 GGA24||LB(CP)
+|-
+|9/26 Ptet+RBS-GFP-DT ||LB(CP)
+|}
+</div>
+incubate 37&deg;C
+===Restriction Enzyme Digestion===
+<div class="experiment">
+<span class="author">Kojima</span>
+{| class="wikitable"
+!||9/24 pSB1C3||EcoRI||SpeI||buffer||MilliQ||total
+|-
+|2cuts||11&micro;L||1&micro;L||1&micro;L||3&micro;L||14&micro;L||30&micro;L
+|-
+|NC||0.5&micro;L||0&micro;L||0&micro;L||1&micro;L||8.5&micro;L||10&micro;L
+|}
+{| class="wikitable"
+!||9/24 Pcon-pT181attenuator-aptamer12-1R-DT||EcoRI||SpeI||buffer||MilliQ||total
+|-
+|2cuts||8.7&micro;L||1&micro;L||1&micro;L||3&micro;L||16.3&micro;L||30&micro;L
+|-
+|NC||0.4&micro;L||0&micro;L||0&micro;L||1&micro;L||8.6&micro;L||10&micro;L
+|}
+{| class="wikitable"
+!||9/24 pT181attenuator||EcoRI||SpeI||buffer||MilliQ||total
+|-
+|2cuts||3.1&micro;L||1&micro;L||1&micro;L||3&micro;L||21.9&micro;L||30&micro;L
+|-
+|NC||0.2&micro;L||0&micro;L||0&micro;L||1&micro;L||8.8&micro;L||10&micro;L
+|}
+{| class="wikitable"
+!||8/17 RBS-GFP-DT||EcoRI||BSA ||buffer||MilliQ||total
+|-
+|1cut||16.4&micro;L||1&micro;L||3&micro;L||3&micro;L||6.6&micro;L||30&micro;L
+|-
+|NC||0.4&micro;L||0&micro;L||1&micro;L||1&micro;L||7.6&micro;L||10&micro;L
+|}
+{| class="wikitable"
+!||9/17 pSB4K5||EcoRI||PstI||BSA ||buffer||MilliQ||total
+|-
+|2cuts||8.5&micro;L||1&micro;L||1&micro;L||3&micro;L||3&micro;L||13.5&micro;L||30&micro;L
+|-
+|NC||0.4&micro;L||0&micro;L||0&micro;L||1&micro;L||1&micro;L||7.6&micro;L||10&micro;L
+|}
+{| class="wikitable"
+!||9/14 pSB4K5||EcoRI||PstI||BSA ||buffer||MilliQ||total
+|-
+|2cuts||7.3&micro;L||1&micro;L||1&micro;L||3&micro;L||3&micro;L||14.7&micro;L||30&micro;L
+|-
+|NC||0.4&micro;L||0&micro;L||0&micro;L||1&micro;L||1&micro;L||7.6&micro;L||10&micro;L
+|}
+{| class="wikitable"
+!||9/22 Pcon-pT181attenuator-DT||EcoRI||SpeI ||buffer||MilliQ||total
+|-
+|2cuts||5.4&micro;L||1&micro;L||1&micro;L||3&micro;L||19.6&micro;L||30&micro;L
+|-
+|NC||0.3&micro;L||0&micro;L||0&micro;L||1&micro;L||8.7&micro;L||10&micro;L
+|}
+{| class="wikitable"
+!||8/7 RBS-GFP-DT 1||XbaI||PstI||BSA ||buffer||MilliQ||total
+|-
+|2cuts||8.1&micro;L||1&micro;L||1&micro;L||3&micro;L||3&micro;L||13.9&micro;L||30&micro;L
+|-
+|NC||0.4&micro;L||0&micro;L||0&micro;L||1&micro;L||1&micro;L||7.6&micro;L||10&micro;L
+|}
+{| class="wikitable"
+!||9/26 RBS-GFP-DT 1||XbaI ||BSA ||buffer||MilliQ||total
+|-
+|1cut||8.1&micro;L||1&micro;L||3&micro;L||3&micro;L||14.9&micro;L||30&micro;L
+|}
+{| class="wikitable"
+!||9/16 P&lambda;-luxI||EcoRI||PstI||BSA ||buffer||MilliQ||total
+|-
+|2cuts||14.7&micro;L||1&micro;L||1&micro;L||3&micro;L||3&micro;L||7.3&micro;L||30&micro;L
+|-
+|NC||0.7&micro;L||0&micro;L||0&micro;L||1&micro;L||1&micro;L||7.3&micro;L||10&micro;L
+|}
+{| class="wikitable"
+!||9/21 Ptet||EcoRI||SpeI ||buffer||MilliQ||total
+|-
+|2cuts||14.2&micro;L||1&micro;L||1&micro;L||3&micro;L||10.8&micro;L||30&micro;L
+|-
+|NC||0.7&micro;L||0&micro;L||0&micro;L||1&micro;L||8.3&micro;L||10&micro;L
+|}
+{| class="wikitable"
+!||9/24 Pcon-pT181attenuator-aptamer12-1R-DT||XbaI||PstI||BSA ||buffer||MilliQ||total
+|-
+|2cuts||8.7&micro;L||1&micro;L||1&micro;L||3&micro;L||3&micro;L||13.3&micro;L||30&micro;L
+|-
+|NC||0.4&micro;L||0&micro;L||0&micro;L||1&micro;L||1&micro;L||7.6&micro;L||10&micro;L
+|}
+</div>
+===Electrophoresis===
+<div class="experiment">
+<span class="author">Kojima</span>
+{| class="wikitable"
+!Lane||Sample|| Enzyme1|| Enzyme2
+|-
+|1||pSB1C3||EcoRI||SpeI
+|-
+|2||pSB1C3||--||--
+|-
+|3||Pcon-pT181attenuator-aptamer12-1R-DT||EcoRI||SpeI
+|-
+|4||Pcon-pT181attenuator-aptamer12-1R-DT||--||--
+|-
+|5||1kb ladder||--||--
+|-
+|6||pT181attenuator||EcoRI||SpeI
+|-
+|7||pT181attenuator||--||--
+|-
+|8||RBS-GFP-DT||EcoRI||--
+|-
+|9||RBS-GFP-DT||--||--
+|}
+[[File:IgkuxxxxBa 1.jpg]]<br>

Template:Kyoto/Notebook/Sep 26

From 2013.igem.org

Revision as of 07:13, 27 September 2013

Contents

Sep 26

Electrophoresis

Ligasion(Golden Gate Assenbly)

Liquid Culture

BSAI Digestion

Colony PCR

Electrophoresis

Electrophoresis

Liquid Culture

Colony PCR

Ligasion(Golden Gate Assenbly)

Plating

Restriction Enzyme Digestion

Electrophoresis