Template:Kyoto/Notebook/Sep 16

From 2013.igem.org

(Difference between revisions)
(→Ligation)
(→Sep 16)
Line 13: Line 13:
===Colony PCR===
===Colony PCR===
-
<!-- ここから -->
 
<div class="experiment">
<div class="experiment">
<span class="author">No name</span>
<span class="author">No name</span>
Line 33: Line 32:
===Gel Extraction===
===Gel Extraction===
<div class="experiment">
<div class="experiment">
-
<span class="author">じっけんしゃ</span>
+
<span class="author">No name</span>
{| class="wikitable"
{| class="wikitable"
!Lane||DNA||Enzyme
!Lane||DNA||Enzyme
Line 39: Line 38:
|1||1kb ladder
|1||1kb ladder
|-
|-
-
|2||pT181 attenuator||X+P
+
|2||pT181 attenuator||XbaI+PstI
|-
|-
-
|3||pT181 attenuator||X+P
+
|3||pT181 attenuator||XbaI+PstI
|-
|-
-
|4||pT181 attenuator||X+P
+
|4||pT181 attenuator||XbaI+PstI
|-
|-
-
|6||pSB1C3||E+S
+
|6||pSB1C3||EcoRI+SpeI
|-
|-
-
|7||pSB1C3||E+S
+
|7||pSB1C3||EcoRI+SpeI
|-
|-
-
|8||pSB1C3||E+S
+
|8||pSB1C3||EcoRI+SpeI
|-
|-
-
|10||pSB4K5||X+P
+
|10||pSB4K5||XbaI+PstI
|-
|-
-
|11||pSB4K5||X+P
+
|11||pSB4K5||XbaI+PstI
|-
|-
-
|12||pSB4K5||X+P
+
|12||pSB4K5||XbaI+PstI
|-
|-
|}
|}
Line 63: Line 62:
!Name||concentration[&micro;g/mL]||260/280||260/230
!Name||concentration[&micro;g/mL]||260/280||260/230
|-
|-
-
|pT181 attenuator(X+P)||8.2||1.62||0.11
+
|pT181 attenuator(XbaI+PstI)||8.2||1.62||0.11
|-
|-
-
|pSB1C3(E+S)||49.5||1.48||0.18
+
|pSB1C3(EcoRI+SpeI)||49.5||1.48||0.18
|-
|-
-
|pSB4K5(X+P)||18.1||1.73||0.57
+
|pSB4K5(XbaI+PstI)||18.1||1.73||0.57
|-
|-
-
|DT(S+P)||70.4||1.36||1.02
+
|DT(SpeI+PstI)||70.4||1.36||1.02
|-
|-
-
|aptamer12-1R-DT(S+P)||42.0||1.82||0.47
+
|aptamer12-1R-DT(SpeI+PstI)||42.0||1.82||0.47
|-
|-
-
|spinach-DT(S+P)||36.8||1.79||0.61
+
|spinach-DT(SpeI+PstI)||36.8||1.79||0.61
|-
|-
|}
|}
Line 102: Line 101:
===PCR===
===PCR===
-
<!-- ここから -->
 
<div class="experiment">
<div class="experiment">
<span class="author">No name</span>
<span class="author">No name</span>
Line 134: Line 132:
|1000pg||2.5||0.5||2.5||2.5||1.5||0.75||0.75||14||25
|1000pg||2.5||0.5||2.5||2.5||1.5||0.75||0.75||14||25
|-
|-
-
|nega||0||0.5||2.5||2.5||1.5||0.75||0.75||16.5||25
+
|NC||0||0.5||2.5||2.5||1.5||0.75||0.75||16.5||25
|}
|}
{| class="wikitable"
{| class="wikitable"
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===Electrophoresis===
===Electrophoresis===
-
 
-
<!-- こっから -->
 
<div class="experiment">
<div class="experiment">
-
<span class="author">じっけんしゃ</span>
+
<span class="author">Yoshida</span>
{| class="wikitable"
{| class="wikitable"
!Lane||Sample
!Lane||Sample
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|1||100bp ladder
|1||100bp ladder
|-
|-
-
|2||anti 1000pg
+
|2||antisense 1000pg
|-
|-
-
|3||anti 800pg
+
|3||antisense 800pg
|-
|-
-
|4||anti 400pg
+
|4||antisense 400pg
|-
|-
|5||tet aptamer-DT 1000pg
|5||tet aptamer-DT 1000pg
Line 166: Line 162:
|7||tet aptamer-DT 400pg
|7||tet aptamer-DT 400pg
|-
|-
-
|8||nega
+
|8||NC
|}
|}
[[File:igku_xxxxxx.xxx]]<br>
[[File:igku_xxxxxx.xxx]]<br>
===Restriction Enzyme Digestion===
===Restriction Enzyme Digestion===
-
<!-- こっから -->
 
<div class="experiment">
<div class="experiment">
-
<span class="author">じっけんしゃ</span>
+
<span class="author">Tatsui</span>
{| class="wikitable"
{| class="wikitable"
-
! ||9/16 Pcon-RBS-tetR-DT(&micro;L)||E(&micro;L)||S(&micro;L)||X(&micro;L)||P(&micro;L)||Buffer(&micro;L)||BSA(&micro;L)||MilliQ(&micro;L)||total(&micro;L)
+
! ||9/16 Pcon-RBS-tetR-DT(&micro;L)||EcoRI(&micro;L)||SpeI(&micro;L)||XbaI(&micro;L)||PstI(&micro;L)||Buffer(&micro;L)||BSA(&micro;L)||MilliQ(&micro;L)||total(&micro;L)
|-
|-
|2 cuts||15.8||0||0||1||1||3||3||6.2||30
|2 cuts||15.8||0||0||1||1||3||3||6.2||30
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|}
|}
{| class="wikitable"
{| class="wikitable"
-
! ||9/16 pSB4K5(&micro;L)||E(&micro;L)||S(&micro;L)||X(&micro;L)||P(&micro;L)||Buffer(&micro;L)||BSA(&micro;L)||MilliQ(&micro;L)||total(&micro;L)
+
! ||9/16 pSB4K5(&micro;L)||EcoRI(&micro;L)||SpeI(&micro;L)||XbaI(&micro;L)||PstI(&micro;L)||Buffer(&micro;L)||BSA(&micro;L)||MilliQ(&micro;L)||total(&micro;L)
|-
|-
|2 cuts||13.5||0||0||1||1||3||3||8.5||30
|2 cuts||13.5||0||0||1||1||3||3||8.5||30
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|}
|}
{| class="wikitable"
{| class="wikitable"
-
! ||8/22 pSB1C3(&micro;L)||E(&micro;L)||S(&micro;L)||X(&micro;L)||P(&micro;L)||Buffer(&micro;L)||BSA(&micro;L)||MilliQ(&micro;L)||total(&micro;L)
+
! ||8/22 pSB1C3(&micro;L)||EcoRI(&micro;L)||SpeI(&micro;L)||XbaI(&micro;L)||PstI(&micro;L)||Buffer(&micro;L)||BSA(&micro;L)||MilliQ(&micro;L)||total(&micro;L)
|-
|-
|2 cuts||18.5||1||0||0||1||3||3||3.5||30
|2 cuts||18.5||1||0||0||1||3||3||3.5||30
Line 196: Line 191:
|}
|}
{| class="wikitable"
{| class="wikitable"
-
! ||8/22 pSB1C3(&micro;L)||E(&micro;L)||S(&micro;L)||X(&micro;L)||P(&micro;L)||Buffer(&micro;L)||BSA(&micro;L)||MilliQ(&micro;L)||total(&micro;L)
+
! ||8/22 pSB1C3(&micro;L)||EcoRI(&micro;L)||SpeI(&micro;L)||XbaI(&micro;L)||PstI(&micro;L)||Buffer(&micro;L)||BSA(&micro;L)||MilliQ(&micro;L)||total(&micro;L)
|-
|-
|2 cuts||18.5||1||1||0||0||3||3||3.5||30
|2 cuts||18.5||1||1||0||0||3||3||3.5||30
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|}
|}
{| class="wikitable"
{| class="wikitable"
-
! ||8/21 pT181 attenuator2(&micro;L)||E(&micro;L)||S(&micro;L)||X(&micro;L)||P(&micro;L)||Buffer(&micro;L)||BSA(&micro;L)||MilliQ(&micro;L)||total(&micro;L)
+
! ||8/21 pT181 attenuator2(&micro;L)||EcoRI(&micro;L)||SpeI(&micro;L)||XbaI(&micro;L)||PstI(&micro;L)||Buffer(&micro;L)||BSA(&micro;L)||MilliQ(&micro;L)||total(&micro;L)
|-
|-
|2 cuts||7.4||1||1||0||0||3||3||14.6||30
|2 cuts||7.4||1||1||0||0||3||3||14.6||30
Line 211: Line 206:
===Ligation===
===Ligation===
-
 
-
<!-- こっから -->
 
<div class="experiment">
<div class="experiment">
-
<span class="author">じっけんしゃ</span>
+
<span class="author">Tatsui</span>
{| class="wikitable"
{| class="wikitable"
!state||colspan="2"|Vector||colspan="2"|Inserter||Ligation High ver.2
!state||colspan="2"|Vector||colspan="2"|Inserter||Ligation High ver.2
|-
|-
-
|experiment||Pcon-pT181anti-spin-DT(X+P)||5.0&micro;g/mL||pSB4K5(X+P)||18.1&micro;g/mL||xx &micro;g/mL
+
|experiment||Pcon-pT181anti-spinach-DT(XbaI+PstI)||5.0&micro;g/mL||pSB4K5(XbaI+PstI)||18.1&micro;g/mL||xx &micro;g/mL
|-
|-
-
|experiment||pC181 attenuator(X+P)||8.2&micro;g/mL||pSB1C3(X+P)||26.0&micro;g/mL||xx &micro;g/mL
+
|experiment||pC181 attenuator(XbaI+PstI)||8.2&micro;g/mL||pSB1C3(XbaI+PstI)||26.0&micro;g/mL||xx &micro;g/mL
|}
|}

Revision as of 18:32, 25 September 2013

Contents

Sep 16

Liquid Culture

No name

Samplemedium
pSB1C3+P-KaiBC①Plusgrow medium(+CP)
pSB1C3+P-KaiBC②Plusgrow medium(+CP)

Colony PCR

No name

Samplebase pair
pSB1C3+P-KaiBC①283
pSB1C3+P-KaiBC②283
PreDenatureDenatureAnnealingExtensioncycle
94°C94°C55°C68°C--
2min30s30s18s30cycles

Gel Extraction

No name

LaneDNAEnzyme
11kb ladder
2pT181 attenuatorXbaI+PstI
3pT181 attenuatorXbaI+PstI
4pT181 attenuatorXbaI+PstI
6pSB1C3EcoRI+SpeI
7pSB1C3EcoRI+SpeI
8pSB1C3EcoRI+SpeI
10pSB4K5XbaI+PstI
11pSB4K5XbaI+PstI
12pSB4K5XbaI+PstI

File:Igku xxbeforexx.xxx
File:Igku xxafterxx.xxx

Nameconcentration[µg/mL]260/280260/230
pT181 attenuator(XbaI+PstI)8.21.620.11
pSB1C3(EcoRI+SpeI)49.51.480.18
pSB4K5(XbaI+PstI)18.11.730.57
DT(SpeI+PstI)70.41.361.02
aptamer12-1R-DT(SpeI+PstI)42.01.820.47
spinach-DT(SpeI+PstI)36.81.790.61

Miniprep

No name

DNAconcentration[µg/mL]260/280260/230
Dx-lux1136.71.461.28
RBS-laczα-DT237.61.881.70
pSB4K5148.41.551.09
Pcon-KBS-tetR-DT126.61.731.36
9/11Plac1160.01.781.28
9/15Plac2144.01.731.81
9/14Plac332.51.512.33
Ptet-RBS-laczα-DT231.31.841.57

PCR

No name

DNA9/6pT181 anti psB1C3(1)KOD plus10x bufferdNTPMgSO4GG1_1C3_fwd primerGG2_1C3_rev primerMilliQtotal
400pg10.52.52.51.50.750.7515.525
800pg20.52.52.51.50.750.7514.525
1000pg2.50.52.52.51.50.750.751425
nega00.52.52.51.50.750.7516.525
PreDenatureDenatureAnnealingExtensioncycle
94°C98°C57°C68°C--
2min10s30s30s30cycles
DNA9/6 12-1R aptamer-DTKOD plus10x bufferdNTPMgSO4GG1_1C3_fwd primerGG1_1C3_rev primerMilliQtotal
400pg10.52.52.51.50.750.7515.525
800pg20.52.52.51.50.750.7514.525
1000pg2.50.52.52.51.50.750.751425
NC00.52.52.51.50.750.7516.525
PreDenatureDenatureAnnealingExtensioncycle
94°C98°C57°C68°C--
2min10s30s30s30cycles

Electrophoresis

Yoshida

LaneSample
1100bp ladder
2antisense 1000pg
3antisense 800pg
4antisense 400pg
5tet aptamer-DT 1000pg
6tet aptamer-DT 800pg
7tet aptamer-DT 400pg
8NC

File:Igku xxxxxx.xxx

Restriction Enzyme Digestion

Tatsui

9/16 Pcon-RBS-tetR-DT(µL)EcoRI(µL)SpeI(µL)XbaI(µL)PstI(µL)Buffer(µL)BSA(µL)MilliQ(µL)total(µL)
2 cuts15.80011336.230
NC0.80000117.210
9/16 pSB4K5(µL)EcoRI(µL)SpeI(µL)XbaI(µL)PstI(µL)Buffer(µL)BSA(µL)MilliQ(µL)total(µL)
2 cuts13.50011338.530
NC0.70000117.310
8/22 pSB1C3(µL)EcoRI(µL)SpeI(µL)XbaI(µL)PstI(µL)Buffer(µL)BSA(µL)MilliQ(µL)total(µL)
2 cuts18.51001333.530
NC0.90000117.110
8/22 pSB1C3(µL)EcoRI(µL)SpeI(µL)XbaI(µL)PstI(µL)Buffer(µL)BSA(µL)MilliQ(µL)total(µL)
2 cuts18.51100333.530
NC0.90000117.110
8/21 pT181 attenuator2(µL)EcoRI(µL)SpeI(µL)XbaI(µL)PstI(µL)Buffer(µL)BSA(µL)MilliQ(µL)total(µL)
2 cuts7.411003314.630
NC0.40000117.610

Ligation

Tatsui

stateVectorInserterLigation High ver.2
experimentPcon-pT181anti-spinach-DT(XbaI+PstI)5.0µg/mLpSB4K5(XbaI+PstI)18.1µg/mLxx µg/mL
experimentpC181 attenuator(XbaI+PstI)8.2µg/mLpSB1C3(XbaI+PstI)26.0µg/mLxx µg/mL
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