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

Line 60: Line 60:
 
<br/>
 
<br/>
 
<h5>Calibration Procedure</h5>
 
<h5>Calibration Procedure</h5>
 +
◻ Added 100 µl LUDOX into wells A1, B1, C1, D1 (or 1 mL LUDOX into cuvette)
 +
<br/>
 +
◻ Added 100 µl of H2O into wells A2, B2, C2, D2 (or 1 mL H2O into cuvette)
 +
<br/>
 +
◻ Measured absorbance 600 nm of all samples in all standard measurement modes in
 +
instrument
 +
<br/>
 +
◻ Recorded the data in the table below or in notebook
 +
<br/>
 +
◻ Imported data into Excel (OD600 reference point tab)​ Sheet_1 provided
 
<br/>
 
<br/>
  

Revision as of 22:09, 29 September 2017

HOME
TEAM
PROJECT
Description
Design
Experiments
InterLab
Model
Results
Attributions
Pictures
HUMAN PRACTICES
Silver HP
Gold HP
AWARDS
Model
Software
SAFETY
JUDGING FORM

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 Eric 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