Difference between revisions of "Team:Florida Atlantic/InterLab"

 
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<img src="https://static.igem.org/mediawiki/2017/archive/2/20/20170929214604%21T--Florida_Atlantic--owlbanner.png">
 
<img src="https://static.igem.org/mediawiki/2017/archive/2/20/20170929214604%21T--Florida_Atlantic--owlbanner.png">
 
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<h1>Interlab Report</h1>
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<h1 style="font-size: 26px">Interlab Report</h1>
  
 
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<center><h4>Member Participations</h4></center>
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<center><h4 style="font-size: 22px">Member Participations</h4></center>
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<h5>Calibrations:</h5>
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<h5 style="font-size: 20px">Calibrations:</h5>
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<p> - Completed by Douglas + Valentina on September 21st</p>
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<p style="font-size: 18px"> - Completed by Douglas + Valentina on September 21st</p>
 
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<br/>
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<h5>Transformations:</h5>
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<h5 style="font-size: 20px">Transformations:</h5>
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<p> - Completed by Douglas + Ariania and supervised by Dr. Pavlovic on September 26th</p>
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<p style="font-size: 18px"> - Completed by Douglas + Ariania and supervised by Dr. Pavlovic on September 26th</p>
 
<br/>
 
<br/>
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<h5>Cell Measurement:</h5>
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<h5 style="font-size: 20px">Cell Measurement:</h5>
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<p> - Completed by Douglas + Rachel S. and assisted by Eric and Daniela (Esiobu lab members) on September 27th, 28th, 29th</p>
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<p style="font-size: 18px"> - Completed by Douglas + Rachel S. and assisted by Erick and Daniela (Esiobu lab members) on September 27th, 28th, 29th</p>
 
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<center><h4>Methodology</h4></center>
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<center><h4 style="font-size: 22px">Methodology</h4></center>
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<h5>Materials</h5>
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<h5 style="font-size: 20px">Materials</h5>
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<p> ● Competent cells (Escherichia coli strain DH5α)  
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<p style="font-size: 18px"> ● Competent cells (Escherichia coli strain DH5α)  
 
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● LB (Luria Bertani) media
 
● LB (Luria Bertani) media
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<br/>
 
<br/>
  
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<h6>Devices (from InterLab Measurement Kit): </h6>
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<h6 style="font-size: 20px">Devices (from InterLab Measurement Kit): </h6>
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<p>● Positive control  
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<p style="font-size: 18px">● Positive control  
 
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<br/>
 
● Negative control  
 
● Negative control  
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<br/>
 
<br/>
 
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<h5>Calibration Procedure</h5>
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<h5 style="font-size: 20px">Calibration Procedure</h5>
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<p>◻ Added 100 µl LUDOX into wells A1, B1, C1, D1 (or 1 mL LUDOX into cuvette)
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<p style="font-size: 18px">◻ Added 100 µl LUDOX into wells A1, B1, C1, D1 (or 1 mL LUDOX into cuvette)
 
<br/>
 
<br/>
 
◻ Added 100 µl of H2O into wells A2, B2, C2, D2 (or 1 mL H2O into cuvette)
 
◻ Added 100 µl of H2O into wells A2, B2, C2, D2 (or 1 mL H2O into cuvette)
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<br/>
 
<br/>
  
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<h5>Transformation Procedure</h5>
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<h5 style="font-size: 20px">Transformation Procedure</h5>
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<p> ◻ added 5uL DNA
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<p style="font-size: 18px"> ◻ added 5uL DNA
 
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◻ let sit 30 min on ice
 
◻ let sit 30 min on ice
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<br/>
 
<br/>
  
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<h5>Cell Measurement Procedure</h5>
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<h5 style="font-size: 20px">Cell Measurement Procedure</h5>
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<h6>Day 1​:</h6>
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</br>
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<p>transform Escherichia coli DH5α with these following plasmids:</p>
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<h6 style="font-size: 19px">Day 1​:</h6>
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<p style="font-size: 18px">transform Escherichia coli DH5α with these following plasmids:</p>
 
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<p>● Positive control
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<p style="font-size: 18px">● Positive control
 
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<br/>
 
● Negative control
 
● Negative control
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● Test Device 6: J23117.BCD2.E0040.B0015</p>
 
● Test Device 6: J23117.BCD2.E0040.B0015</p>
 
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<h6>Day 2​:</h6>
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<h6 style="font-size: 19px">Day 2​:</h6>
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<p> Picked 2 colonies from each of plate and inoculate it on 5-10 mL LB medium + Chloramphenicol.
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<p style="font-size: 18px"> Picked 2 colonies from each of plate and inoculate it on 5-10 mL LB medium + Chloramphenicol.
 
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<br/>
 
Grew the cells overnight (16-18 hours) at 37°C and 220 rpm.
 
Grew the cells overnight (16-18 hours) at 37°C and 220 rpm.
 
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<h6>Day 3​:</h6>
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<h6 style="font-size: 19px">Day 3​:</h6>
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Cell growth, sampling, and assay
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<p style="font-size: 18px"> Cell growth, sampling, and assay
 
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◻ Set instrument to read OD600 (as OD calibration setting)
 
◻ Set instrument to read OD600 (as OD calibration setting)
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◻ Recorded data in notebook</p>
 
◻ Recorded data in notebook</p>
 
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<center><h4>Data</h4></center>
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<center><h4 style="font-size: 22px">Data</h4></center>
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<center>comming soon></center>
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<img style="max-width:95%;border:3px solid darkred;" src="https://static.igem.org/mediawiki/2017/archive/d/d3/20171026205230%21T--Florida_Atlantic--interlabdata.png" width=200; height=700;>
 
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<center><h4>Results</h4></center>
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<center><h4 style="font-size: 22px">Results</h4></center>
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<center><h5>Fluorescence of GFP engineered E.coli Graphs</h5></center>
 
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<center><img src="https://static.igem.org/mediawiki/2017/d/dd/Interlab_Data_Graph.png"></center>
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<center><img style="max-width:95%;border:3px solid darkred;" src="https://static.igem.org/mediawiki/2017/d/dd/Interlab_Data_Graph.png" width=200; height=400;></center>
 
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<center><img src="https://static.igem.org/mediawiki/2017/c/c9/Interlab_Data_Graph2.png"></center>
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<center><img style="max-width:95%;border:3px solid darkred;" src="https://static.igem.org/mediawiki/2017/c/c9/Interlab_Data_Graph2.png" width=200; height=400";></center>
 
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Latest revision as of 20:49, 1 November 2017

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Florida_Atlantic

Interlab Report


Member Participations

Calibrations:

- Completed by Douglas + Valentina on September 21st


Transformations:

- Completed by Douglas + Ariania and supervised by Dr. Pavlovic on September 26th


Cell Measurement:

- Completed by Douglas + Rachel S. and assisted by Erick and Daniela (Esiobu lab members) on September 27th, 28th, 29th



Methodology

Materials

● Competent cells (Escherichia coli strain DH5α)
● LB (Luria Bertani) media
● Chloramphenicol (stock concentration 25 mg/mL dissolved in EtOH - working stock 25 ug/mL)
● 50 ml Falcon tube (or equivalent, preferably amber or covered in foil to block light)
● Incubator at 37°C
● 1.5 ml eppendorf tubes for sample storage
● Ice bucket with ice
● Pipettes
● 96 well plate, 2 different plates used: clear with flat bottom for absorbance; black with flat bottom for fluorescence


Devices (from InterLab Measurement Kit):

● Positive control
● Negative control
● Test Device 1: J23101+I13504
● Test Device 2: J23106+I13504
● Test Device 3: J23117+I13504
● Test Device 4: J23101.BCD2.E0040.B0015
● Test Device 5: J23106.BCD2.E0040.B0015
● Test Device 6: J23117.BCD2.E0040.B0015



Calibration Procedure

◻ Added 100 µl LUDOX into wells A1, B1, C1, D1 (or 1 mL LUDOX into cuvette)
◻ Added 100 µl of H2O into wells A2, B2, C2, D2 (or 1 mL H2O into cuvette)
◻ Measured absorbance 600 nm of all samples in all standard measurement modes in instrument
◻ Recorded the data in the table below or in notebook
◻ Imported data into Excel (OD600 reference point tab)​ Sheet_1 provided


Transformation Procedure

◻ added 5uL DNA
◻ let sit 30 min on ice
◻ heat shocked at 42C for 30 sec
◻ placed back on ice for 2 min
◻ added 450uL Luria Broth and incubated at 37c for 2 hours
◻ plated 100uL on LB/chloramphenicol plates
◻ grew overnight at 37C
◻ this was done for all 8 machines


Cell Measurement Procedure

Day 1​:

transform Escherichia coli DH5α with these following plasmids:


● Positive control
● Negative control
● Test Device 1: J23101+I13504
● Test Device 2: J23106+I13504
● Test Device 3: J23117+I13504
● Test Device 4: J23101.BCD2.E0040.B0015
● Test Device 5: J23106.BCD2.E0040.B0015
● Test Device 6: J23117.BCD2.E0040.B0015


Day 2​:

Picked 2 colonies from each of plate and inoculate it on 5-10 mL LB medium + Chloramphenicol.
Grew the cells overnight (16-18 hours) at 37°C and 220 rpm.

Day 3​:

Cell growth, sampling, and assay
◻ Set instrument to read OD600 (as OD calibration setting)
◻ Measured OD600 of the overnight cultures
◻ Recorded data
◻ Imported data into Excel (Dilution Calculation​) Sheet_1 provided
◻ Diluted the cultures to a target OD600 of 0.02 (see the volume of preloading culture and media in Excel (Dilution Calculation​) Sheet_1) in 15 m​l LB medium + Chloramphenicol in 50 mL falcon tube (amber, or covered with foil to block light).
◻ Incubated the cultures at 37°C and 220 rpm.
◻ Took 1 mL samples of the cultures at 0, 2, 4, and 6 hours of incubation. (At each time point, sampled from each of the 8 devices, two colonies per device, for a total of 16 samples per time point)
◻ Placed samples on ice.
◻ Measured samples (OD and Fl measurement)
◻ Recorded data in notebook


Data


Results


Fluorescence of GFP engineered E.coli Graphs