Template:Kyoto/Notebook/Aug 17

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Contents

Aug 17

Colony PCR

Nakamoto

Samplebase pair
8/15 BBa_K112806(lysis2) -(1)828
negative control--
PreDenatureDenatureAnnealingExtensioncycle
94°C94°C55°C68°C--
5min30s30s1min30cycles

Igku Aug17colonyPCR.jpg

Samplebase pair
8/10 BBa_Q0400 -(1)3995
negative control--
PreDenatureDenatureAnnealingExtensioncycle
94°C94°C55°C68°C--
5min30s30s30s30cycles

File:Image

Liquid Culture

Nakamoto

Samplemedium
8/15 BBa_K112806(lysis2) -(1)Plusgrow medium(+CP)
8/15 BBa_Q04400(tetR) -(1)Plusgrow medium(+Amp)

Master Plate

Nakamoto

NumberUse LB plate (+CP)
1BBa_K112806(lysis2) -(1)
NumberUse LB plate (+Kana)
1BBa_Q04400(tetR) -(1)

Miniprep

Nakamoto and Murakami

DNAconcentration[µg/mL]260/280260/230
K748002(lysis3) -(1)3301.662.08
B0015(DT) -(1)1881.731.82
R0061(Plux) -(1)761.641.30
K117000(lysis1) -(1)2861.712.01
K117000(lysis1) -(2)2481.691.78
I732020(RBS-lacZα-DT) -(1)1981.711.93
I13504(RBS-GFP-DT) -(1)2481.692.03
I13504(RBS-GFP-DT) -(2)2461.551.73
I0462(RBS-luxR-DT) -(1)3261.651.74

Restriction Enzyme Digestion

Murata, Murakami, Nakamoto, Ashida, Matsumoto and Yoshida

8/11 J23100 -(2)SpeIPstIBufferMilliQtotal
2 cuts4.0µL1.0µL1.0µL3.0µL21µL30µL
1 cut1.3µL0.2µL--1.0µL7.5µL10µL
1 cut1.3µL--0.2µL1.0µL7.5µL10µL
NC1.3µL----1.0µL7.7µL10µL
  • at 37°C, for 1h
8/15 B0034(RBS) -(2)SpeIPstIBufferMilliQtotal
2 cuts6.0µL1.0µL1.0µL3.0µL19µL30µL
1 cut2.0µL0.2µL--1.0µL6.8µL10µL
1 cut2.0µL--0.2µL1.0µL6.8µL10µL
NC2.0µL----1.0µL7.0µL10µL
  • at 37°C, for 1h
8/17 K117000(lysis1) -(1)XbaIPstIBSABufferMilliQtotal
2 cuts2.0µL1.0µL1.0µL3.0µL3.0µL20µL30µL
1 cut1.0µL0.2µL--1.0µL1.0µL6.8µL10µL
1 cut1.0µL--0.2µL1.0µL1.0µL6.8µL10µL
NC1.0µL----1.0µL1.0µL7.0µL10µL
  • at 37°C, for 1h
8/17 K748002(lysis3) -(1)XbaIPstIBSABufferMilliQtotal
2 cuts2.0µL1.0µL1.0µL3.0µL3.0µL20µL30µL
1 cut1.0µL0.2µL--1.0µL1.0µL6.8µL10µL
1 cut1.0µL--0.2µL1.0µL1.0µL6.8µL10µL
NC1.0µL----1.0µL1.0µL7.0µL10µL
  • at 37°C, for 1h
8/17 I13504(RBS-GFP-DT) -(1)XbaIPstIBSABufferMilliQtotal
2 cuts2.0µL1.0µL1.0µL3.0µL3.0µL20µL30µL
1 cut1.0µL0.2µL--1.0µL1.0µL6.8µL10µL
1 cut1.0µL--0.2µL1.0µL1.0µL6.8µL10µL
NC1.0µL----1.0µL1.0µL7.0µL10µL
  • at 37°C, for 1h
8/17 I0462(RBS-luxR-DT) -(1)XbaIPstIBSABufferMilliQtotal
2 cuts2.0µL1.0µL1.0µL3.0µL3.0µL20µL30µL
1 cut1.0µL0.2µL--1.0µL1.0µL6.8µL10µL
1 cut1.0µL--0.2µL1.0µL1.0µL6.8µL10µL
NC1.0µL----1.0µL1.0µL7.0µL10µL
  • at 37°C, for 1h
8/17 I732020(RBS-lacZα-DT) -(1)XbaIPstIBSABufferMilliQtotal
2 cuts2.0µL1.0µL1.0µL3.0µL3.0µL20µL30µL
1 cut1.0µL0.2µL--1.0µL1.0µL6.8µL10µL
1 cut1.0µL--0.2µL1.0µL1.0µL6.8µL10µL
NC1.0µL----1.0µL1.0µL7.0µL10µL
  • at 37°C, for 1h
8/11 R6061 -(1)SpeIPstIBufferMilliQtotal
2 cuts8.0µL1.0µL1.0µL3.0µL17µL30µL
1 cut3.0µL0.2µL--1.0µL5.8µL10µL
1 cut3.0µL--0.2µL1.0µL5.8µL10µL
NC3.0µL----1.0µL6.0µL10µL
  • at 37°C, for 1h

Miniprep

Murata

DNAconcentration[µg/mL]260/280260/230
8/17 PBR322(TaKaRa) -(1)471.681.55
8/17 K112806(lysis2) -(1)641.671.70