Laboratory Methods
GST Fusion Protein Purification from Yeast : This protocol has be used to assay binding of Sst2-RGS and Gbeta/gamma to Gpa1-GST.
TAP Fusion Protein Purification from Yeast
FLAG Fusion Protein Purification from Yeast
Liquid Beta-galactosidase Reporter Gene Assay : This protocol is used to quantitatively measure the pheromone response over a range of pheromone concentrations.
Pheromone Halo Assay : This is a bio-assay that measures the responsiveness of cells to a factor pheromone. The assay is easy to conduct and the results are usually unambiguous and highly reproducible.
SDS Whole Cell Extracts : This is a simple and rapid method for extracting total proteins from Yeast for SDS-PAGE and Western Blotting.
TCA Whole Cell Extracts : This is a quantitative method for extracting total proteins for Western Blotting, especially good with phospho-MAPK antibodies.
Sucrose Density Gradient Fractionation of Yeast Membranes : This is an excellent protocol for localization of membrane associated proteins in yeast.
Beta-galactosidase Filter Assay : This method has been used in our lab to screen for and examine mutants with altered pheromone responses.
“Lazy Bones” PLATE Transformation : This is a quick method for plasmid transformation into yeast.
Preparation of Yeast cells for FACS analysis
Plasmid Isolation from Yeast using Qiagen “QiaPrep” Kit
Plasmid Library Amplification : This protocol has been used successfully in our lab to amplify a LEU2 CEN library. After several screens we determined that the quality of the amplified library was excellent.
Paula’s Perfect Immunofluorescence Protocol
Other Protocol Pages: