IGEM:British Columbia/2009/Notebook/Biosensor Sensitivity/2009/05/25: Difference between revisions

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* Will test a different aliquot of chloramphenicol first; if that doesn`t work, then try growing in liquid media.
* Will test a different aliquot of chloramphenicol first; if that doesn`t work, then try growing in liquid media.
* If that works, then next time we make LB-Chlo it will have to be dissolved in the liquid media before it is poured.
* If that works, then next time we make LB-Chlo it will have to be dissolved in the liquid media before it is poured.
* If liquid media doesn't work, then make a new batch of LB-Chlo.
* If liquid media doesn't work, then make a new batch of LB-Chlo plates.


==Antibiotic Liquid Media==
==Antibiotic Liquid Media==

Revision as of 18:27, 25 May 2009

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Antibiotic Plates

Test on Kanamycin Plates

  • Kan plates worked well; growth observed on LB-only but not on LB-Kan.
  • Smear of growth observed on LB-only plate probably due to plates not drying well.

Test on Chloramphenicol plates

  • Chloramphenicol plates did not work well; growth observed on both LB-Chlo plates and LB-only plates.
  • Could be due to aliquot of chloramphenicol not working at the desired concentrations.
  • Will test a different aliquot of chloramphenicol first; if that doesn`t work, then try growing in liquid media.
  • If that works, then next time we make LB-Chlo it will have to be dissolved in the liquid media before it is poured.
  • If liquid media doesn't work, then make a new batch of LB-Chlo plates.

Antibiotic Liquid Media

  • Made LB Broth+Amp (50µL Amp in 50mL LB Broth).
  • Stored on the shelf of Roza's (original) bench.

Transformation of Biobricks

  • The Amp Construction Plasmid plate showed growth of cell colonies
  • The Chloramphenicol Construction Plasmid plate had a smear growth pattern, possibly due to the plates not drying well
  • All other plates had no growth
  • These observed results may have been caused by low plasmid DNA concentrations, will do a test to determine the concentration of the resuspended Biobricks

Test of Resuspended Biobrick Concentrations

Biobrick Part Concentration (µg/µl) A260/280 A260/230
Amp Construction Plasmid 63.96 0.84 1.63
Kan Construction Plasmid 89.89 0.80 1.60
Terminator 63.85 0.81 1.59
J23100/RBS 71.65 1.00 2.10
pBAD 65.47 0.84 1.58
Tet Construction Plasmid 64.60 0.83 1.55
Chlo Construction Plasmid 67.14 0.89 1.72
GFP 62.47 0.84 1.64

Notes

  • 1 ul of resuspended biobrick was used with a Nanodrop 1000 to get the above results
  • The red dye in the biobrick solution may influence the purity readings by shifting the spectrum

Conclusions

  • Low transformation results may not have been due to concentration of DNA, as they were all the same. Nonetheless, we think that for some parts, there may not have been enough DNA, so transformation done tomorrow would be with 2µL of DNA and not 1 µL.

Innoculation of Amp Construction Plasmid Cells

  • Since the Amp construction plasmid cells were successfully transformed, they were innoculated into 10mL of liquid LB+Amp and incubated in the AMBL 37ºC shaking incubator overnight, shaking at 100 rpm.


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