The Thermo Scientific Yeast β-Galactosidase Assay Kit is specially formulated for yeast cells and ideal for identifying protein interactions in vivo using two-hybrid systems.
The Yeast β-Galactosidase Assay Kit allows for qualitative or quantitative determination of β-galactosidase activity in solution directly from colonies growing on solid medium. A portion of the colony is suspended in a mixture of Thermo Scientific Y-PER Yeast Protein Extraction Reagent and the β-Galactosidase Assay Buffer. After a brief incubation period, the solution turns yellow from the hydrolysis of o-nitrophenyl-β-D-galactopyranoside (ONPG) to o-nitrophenol (ONP) and galactose in a mildly alkaline solution. The assay becomes quantitative if calibrated to cell number by measuring the optical density at 660nm.
- Efficient lysis of yeast cells and a colorimetric detection system
- Quantitative or qualitative assay for yeast colonies grown on solid media or in suspension
- Colorimetric assay easily measured in a plate reader or spectrophotometer (420nm)
- Adaptable to 96-well microplates or tube assay formats
- Perform homogeneous screening assays or measure activity secondarily after cell lysis
- Also compatiible with prepared lysates from other species such as bacteria
||Linearity of β-galactosidase assay from cells growing in media in a 96-well plate. Yeast strain BRS1002 carrying plasmid pYX122-β-Gal was grown to an OD600 of 1.0, then 100μL of cells were transferred to a 96-well plate. At time 0, 100μL of a 1:1 mixture of lysis reagent and 2X β-Gal assay buffer were added to each well and absorbance at 420nm was determined. Specific activity for this sample is 160 U (unit = OD405 x 1000/time/OD660).
The gene encoding β-galactosidase (lacZ) of E. coli has been widely used as a reporter gene in many different prokaryotic and eukaryotic organisms. β-Galactosidase activity is also commonly used as an indicator of protein:protein interactions in vivo using two-hybrid systems. The interaction strength is verified or quantitated using a β-galactosidase activity assay.
Common protocols for determining β-galactosidase activity from liquid cultures involve harvesting cells by centrifugation, washing cells several times, and lysing cells by either subjecting them to freeze/thaw cycles or by treating them with detergents or organic solvents. These traditional protocols are not amenable to projects that involve screening cells growing in 96-well plates. The Yeast β-Galactosidase Assay Kit allows cells obtained from solid media or liquid culture to be assayed directly with no harvesting or washing steps.
Mammalian β-Galactosidase Assay Kit
Y-PER Yeast Protein Extraction Reagent