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qPCA: a scalable assay to measure the perturbation of protein-protein interactions in living cells.

Mol Biosyst. 2013; 
Freschi L, Torres-Quiroz F, Dubé AK, Landry CR.
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Catalog Antibody Lysates were immunoprecipitated for 2 h at 4 1C with THEt c-Myc Tag Antibody, mAB, Mouse (GenScript A00704) coupled to Dynabeadss M-280 Tosylactivated (Life Technologies Corporation), washed 3 times with 500 mL of cold washing buffer (0. Get A Quote

摘要

One of the most important challenges in systems biology is to understand how cells respond to genetic and environmental perturbations. Here we show that the yeast DHFR-PCA, coupled with high-resolution growth profiling (DHFR-qPCA), is a straightforward assay to study the modulation of protein-protein interactions (PPIs) in vivo as a response to genetic, metabolic and drug perturbations. Using the canonical Protein Kinase A (PKA) pathway as a test system, we show that changes in PKA activity can be measured in living cells as a modulation of the interaction between its regulatory (Bcy1) and catalytic (Tpk1 and Tpk2) subunits in response to changes in carbon metabolism, caffeine and methyl methanesulfonate (MMS) ... More

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