Template:Kyoto/Notebook/Sep 25

From 2013.igem.org

(Difference between revisions)
(→Gel Extraction)
(→Gel Extraction)
Line 233: Line 233:
[[File:igku_9253.jpg]]<br>
[[File:igku_9253.jpg]]<br>
[[File:igku_9254.jpg]]<br>
[[File:igku_9254.jpg]]<br>
-
{| class="wikitable"
 
-
!Name||concentration[&micro;g/mL]||260/280||260/230
 
-
|-
 
-
|（´ω｀）|| || ||
 
-
|-
 
-
|（ΦωΦ^）|| || ||
 
-
|}
 
-
</div>
 
<!-- ここまでをコピーしてね -->
<!-- ここまでをコピーしてね -->
<br>
<br>
Line 259: Line 251:
[[File:igku_9255.jpg]]<br>
[[File:igku_9255.jpg]]<br>
[[File:igku_9256.jpg]]<br>
[[File:igku_9256.jpg]]<br>
-
{| class="wikitable"
 
-
!Name||concentration[&micro;g/mL]||260/280||260/230
 
-
|-
 
-
|（´ω｀）|| || ||
 
-
|-
 
-
|（ΦωΦ^）|| || ||
 
-
|}
 
{| class="wikitable"
{| class="wikitable"
!Lane||DNA||Enzyme
!Lane||DNA||Enzyme
Line 279: Line 264:
[[File:igku_9257.jpg]]<br>
[[File:igku_9257.jpg]]<br>
[[File:igku_9258.jpg]]<br>
[[File:igku_9258.jpg]]<br>
-
{| class="wikitable"
 
-
!Name||concentration[&micro;g/mL]||260/280||260/230
 
-
|-
 
-
|（´ω｀）|| || ||
 
-
|-
 
-
|（ΦωΦ^）|| || ||
 
-
|}
 
{| class="wikitable"
{| class="wikitable"
!Lane||DNA||Enzyme
!Lane||DNA||Enzyme
Line 299: Line 277:
[[File:igku_9259.jpg]]<br>
[[File:igku_9259.jpg]]<br>
[[File:igku_92510.jpg]]<br>
[[File:igku_92510.jpg]]<br>
-
{| class="wikitable"
 
-
!Name||concentration[&micro;g/mL]||260/280||260/230
 
-
|-
 
-
|（´ω｀）|| || ||
 
-
|-
 
-
|（ΦωΦ^）|| || ||
 
-
|}
 
{| class="wikitable"
{| class="wikitable"
!Lane||DNA||Enzyme
!Lane||DNA||Enzyme
Line 320: Line 291:
[[File:igku_92511.jpg]]<br>
[[File:igku_92511.jpg]]<br>
[[File:igku_92512.jpg]]<br>
[[File:igku_92512.jpg]]<br>
 +
{| class="wikitable"
{| class="wikitable"
!Name||concentration[&micro;g/mL]||260/280||260/230
!Name||concentration[&micro;g/mL]||260/280||260/230
|-
|-
-
|（´ω｀）|| || ||
+
|DT(2-1)||24.9||1.73||0.37
 +
|-
 +
|Ptet(E-0)||12.8||1.59||0.90
 +
|-
 +
|Pcon-antisense(E-2A)||16.5||1.72||1.07
 +
|-
 +
|Pcon-attenuator-aptamer-DT(E-1A)||22.1||1.75||0.94
 +
|-
 +
|Pcon-antisense-spinach-DT(E-1A)||31.0||1.78||1.15
 +
|-
 +
|spinach-DT(1-2)||23.4||1.85||1.11
 +
|-
 +
|Pcon-attenuator(0-1A)||22.5||1.76||0.46
|-
|-
-
|（ΦωΦ^）|| || ||
+
|DT(1-2)||25.1||1.78||1.21
|}
|}

Revision as of 17:33, 26 September 2013

Contents

Sep 25

Electrophoresis

nakamoto

LaneSampleEnzyme1Enzyme2
1100bp ladder
2Pcon-antisense (1C3-66E-fwd A2-662 rev)
3DT (1C3-661-fwd 1C3 662 rev)
4spinach-DT(1C3-661-fwd 1C3 662 rev)
5Pcon-antisense-spinach-DT(1C3-66E fwd A2-661 rev)
6100bp ladder

Igku 9251a.jpg

LaneSampleEnzyme1Enzyme2
2100bp ladder
3Ptet (1C3 66E fwd-1C3 660 rev)
4Pcon-attenuator(1C3-660 fwd A2-661 rev)
5DT(1C3 662 fwd-1C3 661 rev)
6Pcon-attenuator-aptamer-DT(1C3 66E fwd-A2-661-rev)
7100bp ladder

Igku 9251b.jpg

Colony PCR

Nakamoto

Samplebase pair
9/23 Pcon-PT181 attenuator-aptamer12-1R-DT-(5~12)859
9/23 Pcon-PT181 attenuator-RBS-GFP-DT-(5~8)1527
9/23 Pcon-PT181 attenuator-pSB1C3-(5~8)601
PreDenatureDenatureAnnealingExtensioncycle
94 °C94 °C55 °C68 °C--
2 min30 s30 s1min5s30 cycles


Electrophoresis

no name

LaneSampleEnzyme1Enzyme2
1Pcon-PT181 attenuator-aptamer12-1R-DT-5----
2Pcon-PT181 attenuator-aptamer12-1R-DT-6----
3Pcon-PT181 attenuator-aptamer12-1R-DT-7----
4Pcon-PT181 attenuator-aptamer12-1R-DT-8----
5Pcon-PT181 attenuator-aptamer12-1R-DT-9----
6Pcon-PT181 attenuator-aptamer12-1R-DT-10----
7Pcon-PT181 attenuator-aptamer12-1R-DT-11----
8Pcon-PT181 attenuator-aptamer12-1R-DT-12----
91kbp ladder----
10Pcon-PT181 attenuator-RBS-GFP-DT-5----
11Pcon-PT181 attenuator-RBS-GFP-DT-6----
12Pcon-PT181 attenuator-RBS-GFP-DT-7----
13Pcon-PT181 attenuator-RBS-GFP-DT-8----
14Pcon-PT181 attenuator-pSB1C3-5----
15Pcon-PT181 attenuator-pSB1C3-6----
16Pcon-PT181 attenuator-pSB1C3-7----
17Pcon-PT181 attenuator-pSB1C3-8----

Igku 9252.jpg

Transformation

kojima and nakamoto

NameSampleCompetent CellsTotalPlate
Plac+tet aptamer12+Pcon tetR-DT+Ptet+RBS-GFP-DT1µL10µL11µL-
Pcon-pT181attenuator-DT+Pcon-tetR-DT+Ptet+RBS-GFP-DT1µL10µL11µL-
Pcon-tet aptamer12-DT+Pcon-tetR-DT+Ptet+RBS-GFP-DT1µL10µL11µL-
Pcon-Spinach-DT+Pcon-tetR-DT+Ptet+RBS-GFP-DT1µL10µL11µL-
Pcon-pT181antisense+DT+Pcon-tetR-DT+Ptet+RBS-GFP-DT1µL10µL11µL-
Pcon-pT181attenuator+tet aptamer12-DT+Pcon-tetR-DT+Ptet+RBS-GFP-DT1µL10µL11µL-
Pcon tetR-DT+Ptet+RBS-GFP-DT1µL10µL11µL-
Pcon tetR-DT+Pcon-GFP-DT+RBS-GFP-DT1µL10µL11µL-
Pcon-tet aptamer12-DT+Pcon-pT181attenuator+RBS-GFP-DT1µL10µL11µL-
Pcon-pT181attenuator-DT+Pcon-pT181attenuator+RBS-GFP-DT1µL10µL11µL-
Pcon-tet aptamer12-DT+Ptet-pT181antisense+Spinach-DT+Pcon-RBS-GFP-DT+RBS-GFP-DT1µL10µL11µL-
Pcon-tet aptamer12-DT+Pcon-pT181antisense+Spinach-DT+Pcon-pT181attenuator+RBS-GFP-DT1µL10µL11µL-
Pcon-tet aptamer12-DT+Ptet-pT181antisense+Spinach-DT+Pcon-pT181attenuator+RBS-GFP-DT1µL10µL11µL-

NameSampleCompetent CellsTotalPlate
aptamer12-DT(EcoRI+Xbal)+Pcon-pT181attenuator(EcoRI+Spel)2µL20µL22µL-
Ptet(Spel+Pstl)+RBS-GFP-DT(Xbal+Pstl))2µL20µL22µL-

Ligation

nakamoto

stateVectorInserterLigation High ver.2
experiment9/25 aptamer12_1R-DT(EcoRI&XbaI)4.5 µLPcon-pT181 attenuator(EcoRI&SpeI)7.7 µL6.1 µL
experiment9/17 pSB1C3(EcoRI&SpeI)2.0 µL9/3 pT181 attenuator(EcoRI&SpeI)14 µL8 µL
experiment9/25 Pcon-pT181 attenuator(EcoRI&XbaI)4.5 µL9/25 RBS-GFP-DT(EcoRI&SpeI)19.3 µL11.9 µL
experiment9/25 Pcon-RBS-GFP-DT(EcoRI&XbaI)4.2 µLPcon-RBS-tetR-DT(EcoRI&SpeI)20 µL12.1 µL

Restriction Enzyme Digestion

nakamoto>

9/24 Pcon-attenuator-aptamer-DT(µL)E(µL)S(µL)Buffer(µL)MilliQ(µL)total(µL)
2 cuts8.611316.430
NC0.40018.610
9/11 DT(µL)E(µL)X(µL)Buffer(µL)BSA(µL)MilliQ(µL)total(µL)
2 cuts10.4113311.630
NC0.500117.510

Liquid Culture

Kojima

Samplemedium
DT -
pT181attenuator -
Pcon-RBS-tetR-DT -

Gel Extraction

Kojima

LaneDNAEnzyme
1100bp ladder-
2Pcon-pT181antisense
3
4
5-
6DT
7
8
9-
10Spinach-DT
11
12

Igku 9253.jpg
Igku 9254.jpg

LaneDNAEnzyme
1100bp ladder
3Pcon-antisense-spinach-DT(E-1A)
4Pcon-antisense-spinach-DT(E-1A)
5Pcon-antisense-spinach-DT(E-1A)

Igku 9255.jpg
Igku 9256.jpg

LaneDNAEnzyme
7100bp ladder
9Ptet(E-0)
10Ptet(E-0)
11Ptet(E-0)

Igku 9257.jpg
Igku 9258.jpg

LaneDNAEnzyme
13100bp ladder
15Pcon-attenuator(0-1A)
16Pcon-attenuator(0-1A)
17Pcon-attenuator(0-1A)

Igku 9259.jpg
Igku 92510.jpg

LaneDNAEnzyme
19100bp ladder
21Pcon-attenuator-aptamer-DT(E-1A)
22Pcon-attenuator-aptamer-DT(E-1A)
23Pcon-attenuator-aptamer-DT(E-1A)

Igku 92511.jpg
Igku 92512.jpg

Nameconcentration[µg/mL]260/280260/230
DT(2-1)24.91.730.37
Ptet(E-0)12.81.590.90
Pcon-antisense(E-2A)16.51.721.07
Pcon-attenuator-aptamer-DT(E-1A)22.11.750.94
Pcon-antisense-spinach-DT(E-1A)31.01.781.15
spinach-DT(1-2)23.41.851.11
Pcon-attenuator(0-1A)22.51.760.46
DT(1-2)25.11.781.21

Electrophoresis

LaneSampleEnzyme1Enzyme2
11kbp ladder----
2Pcon-pT181 attenuator-aptamer 12_1R-DTES
3Pcon-pT181 attenuator-aptamer 12_1R-DT----
4DTEX
5DT----

Igku 92513.jpg

Gel Extraction

nakamoto

LaneDNAEnzyme
1DTE+X
2DTE+X
3Pcon-pT181 attenuator-aptamer12_1R-DTE+S
4Pcon-pT181 attenuator-aptamer12_1R-DTE+S
5Pcon-pT181 attenuator-aptamer12_1R-DTE+S

Igku 92514.jpg
Igku 92515.jpg

Nameconcentration[µg/mL]260/280260/230
Pcon-pT181 attenuator-aptamer12_1R-DT(E+S)11.81.800.46
DT(E+X)42.61.840.98

Liquid Culture

No name

Samplemedium
Pcon-pT181anntisense-spinach-DT(4k5)-

Ligation

Nakamoto and Tatsui

stateVectorInserterLigation High ver.2
experimentnamexx µLnamexx µLxx µL
NCnamexx µLnamexx µLxx µL

PCR

No name

genome DNAKOD plus10x bufferdNTPMgSO4SasA_fwd primerSasA_rev primerMilliQtotal
0.52.52.51.50.750.75 25
PreDenatureDenatureAnnealingExtensioncycle
　°C　°C　°C　°C--
（◎Д◎）　　　　

Transformation

Tatsui

NameSampleCompetent CellsTotalPlate
aptamer12-1R-DTµLµLµLCP
pSB1C3(EcoRI+Spel)+pT181attenuator(EcoRI+Spal)µLµLµLCP
Pcon-pT181attenuator(EcoRI+Xbal)+RBS-GFP-DT(EcoRI+Spel)µLµLµLAmp
Pcon-RBS-GFP-DT(EcoRI+Xbal)+Pcon-RBS-tetR-DT(EcoRI+Spel)µLµLµLAmp
pSB1C3(EcoRI+Spel)+Pcon-pT181attenuator-aptamer12-1R-DT(EcoRI+Spel)µLµLµLCP

Electrophoresis

Hirano

LaneSampleEnzyme1Enzyme2
1100bp ladder
2DT(2-1)

File:Igku xxxxxx.xxx

精製？

Restriction Enzyme Digestion

DNArestricion enzymerestriction enzymeBufferBSAMilliQtotal
2 cuts（°°） µL µL µL µL µL µL µL
NC µL µL µL µL µL µL µL

液体培養

プレーティング

Colony PCR

No name

Samplebase pair
(｀・ω・´)
PreDenatureDenatureAnnealingExtensioncycle
　°C　°C　°C　°C--
（◎Д◎）　　　　

RNA抽出

cDNA合成

Electrophoresis

Nakamoto

LaneSampleEnzyme1Enzyme2
1100bp ladder
2Pcon-RBS-GFP-DT(RNA)

File:Igku xxxxxx.xxx

Ligation

じっけんしゃ

stateVectorInserterLigation High ver.2
experimentnamexx µLnamexx µLxx µL
NCnamexx µLnamexx µLxx µL

Retrieved from "http://2013.igem.org/Template:Kyoto/Notebook/Sep_25"