Tandem Purification of His6-3x FLAG Tagged Proteins for Mass Spectrometry from Arabidopsis

Abstract

Tandem affinity purification is a powerful method to identify protein complexes that function in association with a known gene of interest. This protocol describes a methodology to capture proteins tagged with His6-3x FLAG explicitly for the purpose of on-bead digestion and identification by mass spectrometry. The high sensitivity and specificity of our methods allow for purification of proteins expressed at native levels from endogenous promoters to enable uncovering the functional roles of plant protein complexes.