Template:Kyoto/Notebook/Sep 6

From 2013.igem.org

Revision as of 18:13, 25 September 2013 by Kojima (Talk | contribs)

Contents

Sep 6

File:Igku Notebook Sep6 1.jpg File:Igku Notebook Sep6 2.jpg File:Igku Notebook Sep6 3.jpg File:Igku Notebook Sep6 4.jpg File:Igku Notebook Sep6 5.jpg File:Igku Notebook Sep6 6.jpg File:Igku Notebook Sep6 7.jpg File:Igku Notebook Sep6 8.jpg File:Igku Notebook Sep6 9.jpg

Electrophoresis

Hirano

LaneSample
1100bp ladder
29/4 RBS-lysis3-DT -3
39/4 RBS-lysis3-DT -4
49/4 RBS-lysis3-DT -5
59/4 RBS-lysis1-DT -3
69/4 Ptrc-KaiC -1
79/4 pSB4K5 -1
8100bp ladder

Igku Sep6 electrophoresis N3.jpg

Colony PCR

No name

Samplebase pair
Pconst-spinach-DT-(1~4)563
Plac-spinach-DT-(1~4)659
Pcon-pT181 antisense-(1~4)468
Ptet-pT181 antisense-(1~4)604
Pconst-pT181 attenuator-(1~4)568
Pconst+RBS-tetR-DT-(1~4)1121
Ptet+RBS-lacZα-DT-(1~4)1162
PreDenatureDenatureAnnealingExtensioncycle
94°C94°C55°C68°C--
5min30s30s40s30cycles

Liquid Culture

No name

Samplemedium
9/4 RBS-Lysis3+DT3Plusgrow medium(+CP)
9/4 RBS-Lysis1+DT3Plusgrow medium(+CP)
9/4 pSB4K5Plusgrow medium(+Kan)

Electrophoresis

No name

LaneSample
1100bp ladder
2Pconst-spinach-DT -(1)
3Pconst-spinach-DT -(2)
4Pconst-spinach-DT -(3)
5Pconst-spinach-DT -(4)
6Plac-spinach-DT -(1)
7Plac-spinach-DT -(2)
8Plac-spinach-DT -(3)
9Plac-spinach-DT -(4)
10Pcon-pT181 antisense -(1)
11Pcon-pT181 antisense -(2)
12Pcon-pT181 antisense -(3)
13Pcon-pT181 antisense -(4)
LaneSample
1100bp ladder
2Ptet-pT181 antisense -(1)
3Ptet-pT181 antisense -(2)
4Ptet-pT181 antisense -(3)
5Ptet-pT181 antisense -(4)
6Plac-pT181 attenuator -(1)
7Plac-pT181 attenuator -(2)
8Plac-pT181 attenuator -(3)
9Plac-pT181 attenuator -(4)

Igku Sep6 electrophosis N4 1.jpg

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 attenuator102.21.982.17
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)XbaIPstIBSABufferMilliQtotal
2 cuts22 µL1 µL1 µL3 µL3 µL0 µL30 µL
NC1.3 µL0 µL0 µL1 µL1 µL6.7 µL10 µL
aptamer-DT (251.7 µg/mL)XbaIPstIBSABufferMilliQtotal
2 cuts0.4 µL1 µL1 µL3 µL3 µL14.1 µL30 µL
NC0.4 µL0 µL0 µL1 µL1 µL7.6 µL10 µL
Pλ-luxI (86.4 µg/mL)EcoRISpeIBSABufferMilliQtotal
2 cuts23.1 µL1 µL1 µL3 µL3 µL1.9 µL30 µL
NC1.2 µL0 µL0 µL1 µL1 µL6.8 µL10 µL
  • at 37 °C, for 1h

Electrophoresis

Hirano

LaneSample
1100bp ladder
2Pconst+RBS-tetR-DT -(1)
3Pconst+RBS-tetR-DT -(2)
4Pconst+RBS-tetR-DT -(3)
5Pconst+RBS-tetR-DT -(4)
6Ptet+RBS-lacZα-DT -(1)
7Ptet+RBS-lacZα-DT -(2)
8100bp ladder
9Ptet+RBS-lacZα-DT -(3)
10Ptet+RBS-lacZα-DT -(4)
11100bp ladder

File:Igku xxxxxxx.xxx

Gel Extraction

No name

LaneDNAEnzyme
1100bp ladder--
2RBS-lysis2-DTXbaI & PstI
3
4----
5aptamer 12_1R-DTXbaI & PstI
6
LaneDNAEnzyme
1100bp ladder--
2Pλ-luxIEcoRI & SpeI
3

File:Igku xxxbeforexxxx.xxx
File:Igku xxxafterxxx.xxx

Nameconcentration[µg/mL]260/280260/230
RBS-lysis2-DT (XbaI & PstI)7.11.880.07
aptamer 12_1R-DT (XbaI & PstI)15.31.930.70
Pλ-luxI (EcoRI & SpeI)4.51.790.31

Liquid Culture

No name

Samplemedium
9/5 Pconst-spinach-DT-(1)Plusgrow medium(+Amp&CP)
9/5 Ptet-pT181 antisense-(1)Plusgrow medium(+CP)
9/5 Pconst+RBS-tetR-DT-(1)Plusgrow medium(+Amp&CP)
9/5 Ptet+RBS-lacZα-DT-(1)Plusgrow medium(+CP)
9/5 Pcon-pT181 attenuator-(1)Plusgrow medium(+CP)
9/4 pT181 antisense-(1)Plusgrow medium(+CP)
9/4 aptamer 12_1R-(1)Plusgrow medium(+CP)

Colony PCR

No name

Samplebase pair
9/5 Plac-spinach-DT-(5~12)563
9/5 Pcon-antisense-(5~12)468
9/5 Plac-attenuator-(5~12)664
9/5 pT181 antisense+Plac-(1)-
PreDenatureDenatureAnnealingExtensioncycle
94°C94°C55°C68°C--
5min30s30s42s30cycles