Team:Aachen/Wetlab/Protocol37

One-step luciferase measurement in yeasts:

Preparation of (1000 ml):


  1. Creation of 5 ml overnight cultures with SC Medium with the desired aminoacids and selections markers. We used a Gal-promoter for the production of the receptors, therefore we used Galactose as carbon source

  2. Dilute the overnights to OD600 0.6 and let the cells grow until OD600 0.8-1.

  3. 90 ul of the cell culture in 96-well microtiterplate and 10ul of the chemical analyte (desired concentration in 10% Ethanol or 1% DMSO)

  4. shake for 20s and incubate by 30ºC during 2.5hours

  5. add 100ul of 10mM Luciferin in a pH of 3 until 5 (diluted in citrat buffer or 1xPBS)

  6. Shake the plate 20s and measure the luminescence within 20seconds.

References

  1. Bioluminescent Yeast Assay for Detection of Organotin Compounds. Grit Kabiersch,Johanna Rajasar̈kka,̈ Marja Tuomela, Annele Hatakka, Marko Virta and Kari Steffen