Difference between revisions of "Team:NTU-Singapore/Measurement"

Line 242: Line 242:
 
<li>Measure OD<sub>600</sub> with 200uL sample</li>
 
<li>Measure OD<sub>600</sub> with 200uL sample</li>
 
<li>Dilute to ODOD<sub>600</sub> = 0.4</li>
 
<li>Dilute to ODOD<sub>600</sub> = 0.4</li>
−
+
<li>Spot 3uL of sample onto LB agar plate</li>
 +
<li>Incubate overnight at 30C</li>
 +
<li>View with DarkReader Transluminator</li>
 
</ol>
 
</ol>
 
</div>
 
</div>

Revision as of 15:47, 18 September 2015

NTU SG iGEM 2015




Measurement

Fluorescence measurement

Material
  • LB
  • MQ water
  • Micro-centrifuge
  • 96-well microplate
  • 50ml Falcon Tube
  • Tecan Infinite m200 microplate reader
  • PBS solution
Procedures
  1. Inoculate strains for overnight culture in 5mL LB
  2. Measure the OD600 of the overnight culture with 200uL of sample
  3. Dilute OD600 to 0.05 in 5mL of LB in 15mL tube
  4. Incubate at 30C for 4 hours so that bacteria reaches mid-log phase
  5. Dilute OD600 to 0.05 in 10mL of LB in 50mL tube
  6. Take 400uL of samples from culture media at one hour intervals starting from T=1hr
  7. Centrifuge at 15000rpm for 3 minutes
  8. Discard supernatant and resuspend pellet with 400uL PBS
  9. Take 150uL of sample and load onto microplate
  10. Measure OD600 and fluorescence intensity


Tecan Infinite M200 Measurement

Material
Parameters
  1. Excitation Wavelength: 485nm
  2. Emission Wavelength: 520nm
  3. Gain: 40


Spotting

Material
  • LB agar plate
  • Microplate Reader
  • LB broth
Parameters
  1. Grow overnight cultures of sample in 5mL LB broth
  2. Measure OD600 with 200uL sample
  3. Dilute to ODOD600 = 0.4
  4. Spot 3uL of sample onto LB agar plate
  5. Incubate overnight at 30C
  6. View with DarkReader Transluminator