Difference between revisions of "Team:CCU Taiwan/Notebook"
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<p class="description">7/3<br> Start interlab experiment-cell measurement<br>7/5<br> YPD formulation<br>7/9<br> Yeast(X33) culture<br>7/13<br> Fundraising briefing session<br>7/16<br> Communicate with NCKU(interlab & | <p class="description">7/3<br> Start interlab experiment-cell measurement<br>7/5<br> YPD formulation<br>7/9<br> Yeast(X33) culture<br>7/13<br> Fundraising briefing session<br>7/16<br> Communicate with NCKU(interlab & | ||
project)<br>7/18<br> Communicate with BIT(project)</p> | project)<br>7/18<br> Communicate with BIT(project)</p> | ||
| − | <p class="description"></p> | + | <p class="description">8/5<br>19:00-20:00<br> Day 2:<br> Pick 2 colonies from each of the transformation plates and inoculate in 5mL LB medium +Chloramphenicol. Grow the cells overnight (14 hours) at 37°C and 220 rpm.</p> |
| − | + | <p class="description">8/6<br>10:00-18:00<br> Day 3: Cell growth, sampling, and assay<br> | |
| + | <ol> | ||
| + | <li>Make a 1:10 dilution of each overnight culture in LB+Chloramphenicol (0.5mL of culture into 4.5mL of LB+Chlor)</li> | ||
| + | <li>Measure Abs600 of these 1:10 diluted cultures</li> | ||
| + | <li>Record the data in your notebook</li> | ||
| + | <li>Dilute the cultures further to a target Abs600 of 0.02 in a final volume of 12 ml LB medium + Chloramphenicol in 50 mL falcon tube (amber, or covered with foil to block light).</li> | ||
| + | <li>Take 500 μL samples of the diluted cultures at 0 hours into 1.5 ml eppendorf tubes, prior to incubation. (At each time point 0 hours and 6 hours, you will take a sample from each of the 8 devices, two colonies per device, for a total of 16 eppendorf tubes with 500 μL samples per time point, 32 samples total). Place the samples on ice.</li> | ||
| + | <li>Measure your samples (Abs600 and Fluorescence measurement)</li> | ||
| + | </ol> | ||
| + | </p> | ||
| + | <br><br> | ||
| + | <p class="description">Digest pGAPZ A(X3)/ pUCIDT_Lac1/ pUCIDT_Px16/ pUCIDT_Px18 with AgeI,EcoRI</P> | ||
| + | <br><br> | ||
| + | <p class="description">8/7<br>10:00-12:00<br>Fluorescence measurement</p> | ||
| + | <p class="description">Ligation</p> | ||
| + | <p class="description">8/8<br> Day 2:<br> Pick 2 colonies from each of the transformation plates and inoculate in 5mL LB medium +Chloramphenicol. Grow the cells overnight (16 hours) at 37°C and 220 rpm.<br>Transformation | ||
| + | <ol> | ||
| + | <li>Add all pGAPZ A_Lac1/ pGAPZ A_Px16/ pGAPZ A_Px18 (15µl) into 20µl ECOS™ 101 Competent Cells [DH5a]</li> | ||
| + | <li>Incubate on ice 5 minutes</li> | ||
| + | <li>Heat shock at 42℃ 45 second</li> | ||
| + | <li>Incubate on ice 5 minutes</li> | ||
| + | <li>Add 140µl LB</li> | ||
| + | <li>Incubate at 37℃ 1hr</li> | ||
| + | <li>Spread on LB+ Zeocin plate</li> | ||
| + | </ol> | ||
| + | </p> | ||
| + | <p class="description">Ligation</p> | ||
</div> | </div> | ||
Revision as of 04:18, 13 September 2018
Notebook
February
Molecular biology class
E-coli genomic DNA preparation
E-coli transformation
March
Instrument operation
April
Project brainstorming-Product Positioning, HGS monomer proportion
May
Culture selection-compare Yeast, E-coli, Acetobacter aceti
Gene design
June
Interlab experiment-Calbration1,2,3
July
7/3
Start interlab experiment-cell measurement
7/5
YPD formulation
7/9
Yeast(X33) culture
7/13
Fundraising briefing session
7/16
Communicate with NCKU(interlab &
project)
7/18
Communicate with BIT(project)
8/5
19:00-20:00
Day 2:
Pick 2 colonies from each of the transformation plates and inoculate in 5mL LB medium +Chloramphenicol. Grow the cells overnight (14 hours) at 37°C and 220 rpm.
8/6
10:00-18:00
Day 3: Cell growth, sampling, and assay
- Make a 1:10 dilution of each overnight culture in LB+Chloramphenicol (0.5mL of culture into 4.5mL of LB+Chlor)
- Measure Abs600 of these 1:10 diluted cultures
- Record the data in your notebook
- Dilute the cultures further to a target Abs600 of 0.02 in a final volume of 12 ml LB medium + Chloramphenicol in 50 mL falcon tube (amber, or covered with foil to block light).
- Take 500 μL samples of the diluted cultures at 0 hours into 1.5 ml eppendorf tubes, prior to incubation. (At each time point 0 hours and 6 hours, you will take a sample from each of the 8 devices, two colonies per device, for a total of 16 eppendorf tubes with 500 μL samples per time point, 32 samples total). Place the samples on ice.
- Measure your samples (Abs600 and Fluorescence measurement)
Digest pGAPZ A(X3)/ pUCIDT_Lac1/ pUCIDT_Px16/ pUCIDT_Px18 with AgeI,EcoRI
8/7
10:00-12:00
Fluorescence measurement
Ligation
8/8
Day 2:
Pick 2 colonies from each of the transformation plates and inoculate in 5mL LB medium +Chloramphenicol. Grow the cells overnight (16 hours) at 37°C and 220 rpm.
Transformation
- Add all pGAPZ A_Lac1/ pGAPZ A_Px16/ pGAPZ A_Px18 (15µl) into 20µl ECOS™ 101 Competent Cells [DH5a]
- Incubate on ice 5 minutes
- Heat shock at 42℃ 45 second
- Incubate on ice 5 minutes
- Add 140µl LB
- Incubate at 37℃ 1hr
- Spread on LB+ Zeocin plate
Ligation