Team:Gifu/Notebook
From 2014.igem.org
(Difference between revisions)
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<tr align="center"><td>Mon</td><td>Tue</td><td>Wed</td><td>Thr</td><td>Fri</td><td><font color="blue">Sat</font></td><td><font color="red">Sun</td><td colspan="3"> </td><td>Mon</td><td>Tue</td><td>Wed</td><td>Thr</td><td>Fri</td><td><font color="blue">Sat</font></td><td><font color="red">Sun</td></tr> | <tr align="center"><td>Mon</td><td>Tue</td><td>Wed</td><td>Thr</td><td>Fri</td><td><font color="blue">Sat</font></td><td><font color="red">Sun</td><td colspan="3"> </td><td>Mon</td><td>Tue</td><td>Wed</td><td>Thr</td><td>Fri</td><td><font color="blue">Sat</font></td><td><font color="red">Sun</td></tr> | ||
<tr align="center"><td></td><td></td><td></td><td></td><td>1</td><td><font color="blue">2</td><td><font color="red">3</td> | <tr align="center"><td></td><td></td><td></td><td></td><td>1</td><td><font color="blue">2</td><td><font color="red">3</td> | ||
- | <td colspan="3"> </td><td><a href="#0901" style="color: black ;"><u>1</td><td | + | <td colspan="3"> </td><td><a href="#0901" style="color: black ;"><u>1</td><td>2</td><td><a href="#0903" style="color: black ;"><u>3</td><td><a href="#0904" style="color: black ;"><u>4</td><td><a href="#0905" style="color: black ;"><u>5</td><td><font color="blue">6</td><td><font color="red">7</td></tr> |
<tr align="center"><td>4</td><td>5</td><td>6</td><td>7</td><td>8</td><td><font color="blue">9</td><td><font color="red">10</td> | <tr align="center"><td>4</td><td>5</td><td>6</td><td>7</td><td>8</td><td><font color="blue">9</td><td><font color="red">10</td> | ||
<td colspan="3"> </td><td><a href="#0908" style="color: black ;"><u>8</td><td><a href="#0909" style="color: black ;"><u>9</td><td><a href="#0910" style="color: black ;"><u>10</td><td><a href="#0911" style="color: black ;"><u>11</td><td><a href="#0912" style="color: black ;"><u>12</td><td><font color="blue"><a href="#0913" style="color: blue ;"><u>13</td><td><font color="blue"><a href="#0914" style="color: red ;"><u>14</td></tr> | <td colspan="3"> </td><td><a href="#0908" style="color: black ;"><u>8</td><td><a href="#0909" style="color: black ;"><u>9</td><td><a href="#0910" style="color: black ;"><u>10</td><td><a href="#0911" style="color: black ;"><u>11</td><td><a href="#0912" style="color: black ;"><u>12</td><td><font color="blue"><a href="#0913" style="color: blue ;"><u>13</td><td><font color="blue"><a href="#0914" style="color: red ;"><u>14</td></tr> | ||
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<p align="right"><a href="#calender">back to calender</a></p> | <p align="right"><a href="#calender">back to calender</a></p> | ||
- | + | <h2><a name="0903"></a>September 3rd</h2> | |
- | <h2><a name=""></a></h2> | + | <h3>Culture</h3> |
- | <h3></h3> | + | <ol class="list"> |
- | < | + | <li>-Lac-3'intron-RBS-RFP(without stop codon)-5'intron M-DT-Cm- 20 mL</li> |
- | < | + | <li>-Lac-3'intron-RBS-RFP(with stop codon)-5'intron M-DT-Cm- 5 mL</li> |
+ | </ol> | ||
<p align="right"><a href="#calender">back to calender</a></p> | <p align="right"><a href="#calender">back to calender</a></p> | ||
- | + | ||
- | + | <h2><a name="0904"></a>September 4th</h2> | |
- | <h2><a name=""></a></h2> | + | <h3>Make circular RNA</h3> |
- | <h3></h3> | + | <ol class="list"> |
- | < | + | <li>TTHA0715</li> |
- | <h3></h3> | + | <li>RFP(without stop codon)</li> |
+ | <li>RFP(with stop codon)</li> | ||
+ | </ol> | ||
+ | <h3>Cell body Crush</h3> | ||
+ | <ol class="list"> | ||
+ | <li>TTHA0715</li> | ||
+ | <li>RFP(without stop codon)</li> | ||
+ | <li>RFP(with stop codon)</li> | ||
+ | </ol> | ||
<p align="right"><a href="#calender">back to calender</a></p> | <p align="right"><a href="#calender">back to calender</a></p> | ||
- | + | ||
- | + | <h2><a name="0905"></a>September 5th</h2> | |
- | <h2><a name=""></a></h2> | + | <h3>SDS-PAGE</h3> |
- | <h3></h3> | + | |
<h3></h3> | <h3></h3> | ||
<h3></h3> | <h3></h3> | ||
<p align="right"><a href="#calender">back to calender</a></p> | <p align="right"><a href="#calender">back to calender</a></p> | ||
- | + | ||
- | + | <h2><a name="0908"></a>September 8th</h2> | |
- | <h2><a name=""></a></h2> | + | |
<h3></h3> | <h3></h3> | ||
<h3></h3> | <h3></h3> | ||
<h3></h3> | <h3></h3> | ||
<p align="right"><a href="#calender">back to calender</a></p> | <p align="right"><a href="#calender">back to calender</a></p> | ||
- | + | ||
<!-- | <!-- | ||
<h2><a name=""></a></h2> | <h2><a name=""></a></h2> |
Revision as of 02:29, 9 September 2014
Calender
Click on a date to see our research activities on that day.
August | September | |||||||||||||||
Mon | Tue | Wed | Thr | Fri | Sat | Sun | Mon | Tue | Wed | Thr | Fri | Sat | Sun | |||
1 | 2 | 3 | 1 | 2 | 3 | 4 | 5 | 6 | 7 | |||||||
4 | 5 | 6 | 7 | 8 | 9 | 10 | 8 | 9 | 10 | 11 | 12 | 13 | 14 | |||
11 | 12 | 13 | 14 | 15 | 16 | 17 | 15 | 16 | 17 | 18 | 19 | 20 | 21 | |||
18 | 19 | 20 | 21 | 22 | 23 | 24 | 22 | 23 | 24 | 25 | 26 | 27 | 28 | |||
25 | 26 | 27 | 28 | 29 | 30 | 31 | 29 | 30 |
Glossary
- His
- alphabet
- RFP
- red fluorescent protein
- 5'-intron
- 5'-intron M
- 3'-intron
- Lac
- Amp
- ampicillin resistance gene
- Cm
- chloramphenicol resistance gene
- RBS
- DT
- EcoRI
- restriction site that is cut by EcoRI or restriction enzyme
- XbaI
- restriction site that is cut by XbaI or restriction enzyme
- SpeI
- restriction site that is cut by SpeI or restriction enzyme
- PstI
- restriction site that is cut by PstI or restriction enzyme
- TTHA0715
- cold shock protein with amino acid sequence cut by thrombin
August
August 11th
Restriction Enzyme Digestion
sample | restriction enzymes |
---|---|
His-RFP(with stop codon) | SpeI,EcoRI | 5'-intron M | SpeI,EcoRI |
Lac-3'intron-RBS | SpeI,EcoRI |
His-RFP(without stop codon) | XbaI,EcoRI |
RBS-Amp | SpeI,EcoRI |
Ligation
- EcoRI-His-RFP(with stop codon)-SpeI + XbaI-DT-Cm-EcoRI
- EcoRI-5'intron M-SpeI + XbaI-DT-Cm-EcoRI
- EcoRI-Lac-3'intron-RBS-SpeI + XbaI-His-RFP(without stop codon)-5'intron-DT-Cm-EcoRI
- EcoRI-His-RFP(without stop codon)-SpeI + SpeI-Amp-EcoRI
Transformation
- -His-RFP(with stop codon)-DT-Cm-
- -5'intron M-DT-Cm-
- -Lac-3'intron-RBS-His-RFP(without stop codon)-5'intron-DT-Cm-
- -His-RFP(without stop codon)-Amp-
Densitometry
sample | concentration(ng/µL) |
---|---|
His-RFP(with stop codon) EcoRI-SpeI | 162.6 |
5'intron M EcoRI-SpeI | 45.4 |
Lac-3'intron-RBS EcoRI-SpeI | 120.9 |
His-RFP(without stop codon) EcoRI-SpeI | 160.0 |
His-RFP(without stop codon)-5'intron-DT-Cm XbaI-EcoRI | 174.0 |
Amp EcoRI-SpeI | 101.8 |
August 12th
Colony PCR
- His-RFP(with stop codon)-DT-Cm
- 5'intron M-DT-Cm
- Lac-3'intron-RBS-His-RFP(without stop codon)-5'intron-DT-Cm:[1]turning red
- Lac-3'intron-RBS-His-RFP(without stop codon)-5'intron-DT-Cm:[2]turning pink
- Lac-3'intron-RBS-His-RFP(without stop codon)-5'intron-DT-Cm:[3]turning white
- His-RFP(without stop codon)-Amp
Electrophoresis
- His-RFP(with stop codon)-DT
- His-RFP
- 5'intron M-DT
- 5'intron M
- Lac-3'intron-RBS-His-RFP(without stop codon)-5'intron-DT:[1]
- Lac-3'intron-RBS-His-RFP(without stop codon)-5'intron-DT:[2]
- Lac-3'intron-RBS-His-RFP(without stop codon)-5'intron-DT:[3]
- His-RFP(without stop codon)-5'intron-DT
- His-RFP(without stop codon)
Liquid Culture
- His-RFP(with stop codon)-DT
- 5'intron M-DT
- Lac-3'intron-RBS-His-RFP(without stop codon)-5'intron-DT:[1]
- Lac-3'intron-RBS-His-RFP(without stop codon)-5'intron-DT:[3]
- His-RFP(without stop codon)
August 13th
Miniprep
PCR
By use of KAPA HiFi DNA Polymerase
- His-RFP(with stop codon)-DT-Cm
- 5'intron M-DT-Cm
By use of Taq DNA Polymerase
- Lac-3'intron-RBS-His-RFP(without stop codon)-5'intron-DT-Cm:[1]turning red
- Lac-3'intron-RBS-His-RFP(without stop codon)-5'intron-DT-Cm:[3]turning white
Electrophoresis
- His-RFP(with stop codon)-DT-Cm
- 5'intron M-DT-Cm
- DNA Ladder Markers DNA Ladder One(Broad Range)
- Lac-3'intron-RBS-His-RFP(without stop codon)-5'intron-DT-Cm:[1]turning red
- Lac-3'intron-RBS-His-RFP(without stop codon)-5'intron-DT-Cm:[3]turning white
Restriction Enzyme Digestion
sample | restriction enzymes |
---|---|
His-RFP(with stop codon)-DT-Cm | EcoRI,XbaI |
5'intron M-DT-Cm | EcoRI,XbaI |
Densitometry
sample | concentration(ng/µL) |
---|---|
EcoRI-Lac-RBS-SpeI | 65.0 |
XbaI-His-RFP(with stop codon)-DT-Cm-EcoRI | 188.8 |
EcoRI-His-RFP(with stop codon)-SpeI | 160.0 |
EcoRI-His-RFP(without stop codon)-SpeI | 160.0 |
XbaI-5'intron M-DT-Cm-EcoRI | 167.8 |
Ligation
- EcoRI-Lac-RBS-SpeI + XbaI-His-RFP(with stop codon)-DT-Cm-EcoRI
- EcoRI-His-RFP(with stop codon)-SpeI + XbaI-5'intron M-DT-Cm-EcoRI
- EcoRI-His-RFP(without stop codon)-SpeI + XbaI-5'intron M-DT-Cm-EcoRI
Transformation
- -Lac-RBS-His-RFP(with stop codon)-DT-Cm-
- -His-RFP(with stop codon)-5'intron M-DT-Cm-
- -His-RFP(without stop codon)-5'intron M-DT-Cm-
August 16th
Liquid Culture
August 18th
Plasmid Extraction
- -Lac-RBS-His-RFP(with stop codon)-DT-Cm-
- -His-RFP(with stop codon)-5'intron M-DT-Cm-
- -His-RFP(without stop codon)-5'intron M-DT-Cm-
Plasmid Purification
- -5'intron M-His-RFP(with stop codon)-
- -5'intron M-His-RFP(without stop codon)-
Miniprep
Colony PCR
Restriction Enzyme Digestion
sample | restriction enzymes |
---|---|
-His-RFP(with stop codon)-5'intron M-DT-Cm- | EcoRI,XbaI |
-His-RFP(without stop codon)-5'intron M-DT-Cm- | EcoRI,XbaI |
Densitometry
sample | concentration(ng/µL) |
---|---|
EcoRI-Lac-3'intron-RBS-SpeI | 120.0 |
XbaI-His-RFP(with stop codon)-5'intron M-DT-Cm-EcoRI | 187.8 |
XbaI-His-RFP(without stop codon)-5'intron M-DT-Cm-EcoRI | 158.9 |
Ligation&Transformation
- EcoRI-Lac-3'intron-RBS-SpeI + XbaI-His-RFP(with stop codon)-5'intron M-DT-Cm-EcoRI
- EcoRI-Lac-3'intron-RBS-SpeI + XbaI-His-RFP(without stop codon)-5'intron M-DT-Cm-EcoRI
Polyethylene glycol Precipitation
August 19th
Colony PCR
- -Lac-3'intron-RBS-His-RFP(with stop codon)-5'intron M-DT-Cm-
- -Lac-3'intron-RBS-His-RFP(without stop codon)-5'intron M-DT-Cm-
Electrophoresis
- -Lac-3'intron-RBS-His-RFP(with stop codon)-5'intron M-DT-Cm-
- -Lac-3'intron-RBS-His-RFP(without stop codon)-5'intron M-DT-Cm-
- marker
- -His-RFP(with stop codon)-5'intron M-DT-Cm-
- -His-RFP(without stop codon)-5'intron M-DT-Cm-
Shaking Culture
August 20th
Colony PCR
- -Lac-3'intron-RBS-His-RFP(without stop codon)-5'intron M-DT-Cm-
by use of another colony differs from 8/19
Electrophoresis
- -Lac-3'intron-RBS-His-RFP(without stop codon)-5'intron M-DT-Cm-
- marker
- -His-RFP(without stop codon)-5'intron M-DT-Cm-
August 21th
Reagent Preparation
Protein Extraction
SDS-PAGE
August 22th
Prorein Purification by use of Ni-NTA
SDS-PAGE
August 23th
Prorein Purification by use of Ni-NTA
SDS-PAGE
August 24th
Extraction of plasmid device
- -Lac-RBS-His-RFP(with stop codon)-DT-Cm-
- -Lac-RBS-3'intron-His-RFP(without stop codon)-5'intron-DT-Cm-
- -Lac-RBS-3'intron-His-RFP(with stop codon)-5'intron M-DT-Cm-
- -Lac-RBS-3'intron-His-RFP(without stop codon)-5'intron M-DT-Cm-
PCR&Electrophoresis
- EcoRI-His-RFP(with stop codon)-PstI
- EcoRI-His-RFP(without stop codon)-PstI
- EcoRI-5'intron M-PstI
- EcoRI-Lac-3'intron-RBS-PstI
August 25th
Extraction of plasmid device
SDS-PAGE
- Lac-3'intron-RBS-His-RFP-5'intron M-DT-Cm
August 26th
PCR
Restriction Enzyme Digestion
sample | restriction enzymes |
---|---|
His-RFP(with stop codon) | EcoRI,PstI |
His-RFP(without stop codon) | EcoRI,PstI |
5'intron M | EcoRI,PstI |
Lac-3'intron-RBS | EcoRI,PstI |
TTHA0175 | EcoRI,SpeI |
Densitometry
sample | concentration(ng/µL) |
---|---|
His-RFP(with stop codon) | 180.7 |
His-RFP(without stop codon) | 164.6 |
5'intron M | 37.7 |
Lac-3'intron-RBS | 107.5 |
TTHA0175 | 112.7 |
Ligation&Transformation
- EcoRI-His-RFP(with stop codon)-PstI + EcoRI-Cm-PstI
- EcoRI-His-RFP(without stop codon)-PstI + EcoRI-Cm-PstI
- EcoRI-5'intron M-PstI + EcoRI-Cm-PstI
- EcoRI-Lac-3'intron-RBS-PstI + EcoRI-Cm-PstI
- EcoRI-TTHA0175-SpeI + EcoRI-5'intron M-DT-Cm-XbaI
August 27th
Colony PCR
- -His-RFP(with stop codon)-Cm-
- -TTHA0175-Amp-
- -TTHA0175-5'intron M-DT-Cm-
Shaking Culture
- -His-RFP(with stop codon)-Cm-
- -TTHA0175-Amp-
- -TTHA0175-5'intron M-DT-Cm-
Enrichment Culture
August 28th
Plasmid Purification
- -His-RFP(with stop codon)-5'intron M-Cm-
- -TTHA0175-5'intron M-DT-Cm-
PCR
- -His-RFP(with stop codon)-Cm-
- -Lac-3'intron-RBS-Cm-
- -His-RFP(without stop codon)-Cm-
- -5'intron M-Cm-
August 29th
Densitometry
sample | concentration(ng/µL) |
---|---|
EcoRI-TTHA0175-5'intron M-DT-Cm-XbaI | 118.8 |
Ligation
- EcoRI-TTHA0175-5'intron M-DT-Cm-XbaI + EcoRI-Lac-3'intron-RBS-SpeI
SDS-PAGE
Electrophoresis
- EcoRI-TTHA0175-5'intron M-DT-Cm-XbaI
August 30th
Colony PCR
- -Lac-3'intron-RBS-TTHA0715-5'intron M-DT-Cm-
- Plate4 3G
August 31th
Colony PCR
Because result of Colony PCR on 8/30 is not good, We chose 5 colonies and put them on PCR.
September 1st
Shaking Culture
- -Lac-3'intron-RBS-TTHA0715-5'intron M-DT-Cm-
Cell body Crush&SDS-PAGE(gel 10%)
September 3rd
Culture
- -Lac-3'intron-RBS-RFP(without stop codon)-5'intron M-DT-Cm- 20 mL
- -Lac-3'intron-RBS-RFP(with stop codon)-5'intron M-DT-Cm- 5 mL
September 4th
Make circular RNA
- TTHA0715
- RFP(without stop codon)
- RFP(with stop codon)
Cell body Crush
- TTHA0715
- RFP(without stop codon)
- RFP(with stop codon)