Template:Kyoto/Notebook/Sep 6

From 2013.igem.org

(Difference between revisions)
(→Miniprep)
(→Colony PCR)
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===Colony PCR===
===Colony PCR===
<div class="experiment">
<div class="experiment">
-
<span class="author">No name</span>
+
<span class="author">Hirano</span>
{| class="wikitable"
{| class="wikitable"
!|Sample||base pair
!|Sample||base pair
|-
|-
-
|8/27 Pcon-pT181 antisense-(1)||542
+
|Pconst＋RBS-tetR-DT-(1)||1121
|-
|-
-
|8/26 pT181 attenuator-DT-(1)||738
+
|Pconst＋RBS-tetR-DT-(2)||1121
-
|}
+
|-
-
{| class="wikitable"
+
|Pconst＋RBS-tetR-DT-(3)||1121
-
!PreDenature||Denature||Annealing||Extension||cycle
+
|-
 +
|Pconst＋RBS-tetR-DT-(4)||1121
 +
|-
 +
|Ptet＋RBS-lacZα-DT-(1)||1162
 +
|-
 +
|Ptet＋RBS-lacZα-DT-(2)||1162
|-
|-
-
|94&deg;C||94&deg;C||55&deg;C||68&deg;C||--
+
|Ptet＋RBS-lacZα-DT-(3)||1162
|-
|-
-
|5min||30s||30s||30s||30cycles
+
|Ptet＋RBS-lacZα-DT-(4)||1162
|}
|}
-
[[File:igku_Aug19electrophoresis1.png]]
 
</div>
</div>

Revision as of 10:04, 24 September 2013

Contents

Sep 6

Liquid Culture

No name

Samplemedium
9/4 RBS-Lysis3＋DT3Plusgrow medium(+Cp)
9/4 RBS-Lysis1＋DT3Plusgrow medium(+Cp)
9/4 PSB4K5Plusgrow medium(+K)　

Colony PCR

No name

Samplebase pair
Pconst-spinach-DT-(1)-
Pconst-spinach-DT-(2)-
Pconst-spinach-DT-(3)-
Pconst-spinach-DT-(4)-
Plac-spinach-DT-(1)-
Plac-spinach-DT-(2)-
Plac-spinach-DT-(3)-
Plac-spinach-DT-(4)-
Pcon-PT181 antisense-(1)-
Pcon-PT181 antisense-(2)-
Pcon-PT181 antisense-(3)-
Pcon-PT181 antisense-(4)-
Samplebase pair
Ptet-PT181 antisense-(1)-
Ptet-PT181 antisense-(2)-
Ptet-PT181 antisense-(3)-
Ptet-PT181 antisense-(4)-
Plac-PT181 attenuater-(1)-
Plac-PT181 attenuater-(2)-
Plac-PT181 attenuater-(3)-
Plac-PT181 attenuater-(4)-

Miniprep

Honda and Kojima

DNAconcentration [µg/mL]260/280260/230
PT181 antisense-(1)238.71.681.44
PT181 antisense-(2)248.81.831.77
PT181 antisense-(3)223.21.781.61
PT181 antisense-(4)220.81.891.92
tetR aptamer-(1)252.51.891.90
tetR aptamer-(2)211.21.901.99
DT181 attenuater102.21.982.17
121R aptamer-DT251.71.922.01
PSB4K5212.91.801.90
RBS-lacZα-DT185.71.781.79
RBS-lysis2-DT-(1)84.81.852.24

Restriction Enzyme Digestion

Tatsui

9/6 RBS-lysis2-DT (78.3 µg/mL)EcoRISpeIXbaIPstIBSABufferMilliQtotal
2 cuts22µL0µL0µL1µL1µL3µL3µL0µL30µL
NC1.3µL0µL0µL0µL0µL1µL1µL6.7µL10µL
  • at 37°C, for 1h
aptamer-DT (251.7 µg/mL)EcoRISpeIXbaIPstIBSABufferMilliQtotal
2 cuts0.4µL0µL0µL1µL1µL3µL3µL14.1µL30µL
NC0.4µL0µL0µL0µL0µL1µL1µL7.6µL10µL
  • at 37°C, for 1h
Pλ-luxI (86.4 µg/mL)EcoRISpeIXbaIPstIBSABufferMilliQtotal
2 cuts23.1µL1µL1µL0µL0µL3µL3µL1.9µL30µL
NC1.2µL0µL0µL0µL0µL1µL1µL6.8µL10µL
  • at 37°C, for 1h

Colony PCR

Hirano

Samplebase pair
Pconst＋RBS-tetR-DT-(1)1121
Pconst＋RBS-tetR-DT-(2)1121
Pconst＋RBS-tetR-DT-(3)1121
Pconst＋RBS-tetR-DT-(4)1121
Ptet＋RBS-lacZα-DT-(1)1162
Ptet＋RBS-lacZα-DT-(2)1162
Ptet＋RBS-lacZα-DT-(3)1162
Ptet＋RBS-lacZα-DT-(4)1162

Gel Extraction

No name

LaneDNAEnzyme
1100bp ladder--
28/20 RBS-lysis1-(1)EcoRI & SpeI
3
4----
58/28 RBS-GFP-DT-(2)XbaI & PstI
6

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

LaneDNAEnzyme
1100bp ladder--
28/28 PluxSpeI & PstI
3
4----
58/20 RBS-tetR-DT-(1)XbaI & PstI
6

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

Nameconcentration[µg/mL]260/280260/230
Pcon-RBS-GFP-DT (EcoRI&SpeI)7.11.880.07
Pcon-RBS-luxR-DT (EcoRI&XbaI)15.31.930.70
Plux (SpeI&PstI)4.51.790.31
RBS-GFP-DT (XbaI&PstI)6.02.160.03
RBS-tetR-DT (XbaI&PstI)7.61.910.38


Liquid Culture

No name

Samplemedium
8/21 RBS-Lysis2-9Plusgrow medium(+Amp)
8/21 RBS-Lysis2-12Plusgrow medium(+Amp)
8/21 RBS-Lysis2-14Plusgrow medium(+Amp)　

Colony PCR

No name

Samplebase pair
8/27 Pcon-pT181 antisense-(1)542
8/26 pT181 attenuator-DT-(1)738
PreDenatureDenatureAnnealingExtensioncycle
94°C94°C55°C68°C--
5min30s30s30s30cycles

File:Igku Aug19electrophoresis1.png

Colony PCR

No name

Samplebase pair
8/27 Pcon-pT181 antisense-(1)542
8/26 pT181 attenuator-DT-(1)738
PreDenatureDenatureAnnealingExtensioncycle
94°C94°C55°C68°C--
5min30s30s30s30cycles

File:Igku Aug19electrophoresis1.png

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