Protein and antibody purification followed by immunoprecipitation of MYB and GATA zinc finger-type maize proteins with magnetic beads

• Published in: STAR protocols Vol. 3; no. 2; p. 101449
• Main Authors: Vélez-Bermúdez, Isabel Cristina, Salazar-Henao, Jorge Enrique, Riera, Marta, Caparros-Ruiz, David, Schmidt, Wolfgang
• Format: Journal Article
• Language: English
• Published: United States Elsevier Inc 17.06.2022; Elsevier
• Subjects: Immunoprecipitation; Magnetic Phenomena; Molecular Biology; Plant sciences; Protein Biochemistry; Protein expression and purification; Proteins - chemistry; Protocol; Zea mays - genetics; Zinc Fingers; Protein Biochemistry; Plant sciences; Molecular Biology; Protein expression and purification
• ISSN: 2666-1667; 2666-1667
• DOI: 10.1016/j.xpro.2022.101449

Co-immunoprecipitation (Co-IP) is a widely used and powerful approach for studying protein-protein interactions in vivo. Here, we describe a protocol for antibody purification and immobilization followed by immunoprecipitation from plant tissue extracts using magnetic beads. The protocol has been used to detect regulators in the Zea mays phenylpropanoid pathway. The protocol is amenable to a variety of downstream assays, including western blotting and mass spectrometry. For complete details on the use and execution of this protocol, please refer to Vélez-Bermúdez et al. (2015). [Display omitted] •Protein expression and purification of MBP fusions•Affinity purification of antibodies•Immunoprecipitation from maize cell extracts with magnetic beads•Amenable to downstream assays like mass spec and western blot Publisher’s note: Undertaking any experimental protocol requires adherence to local institutional guidelines for laboratory safety and ethics. Co-immunoprecipitation (Co-IP) is a widely used and powerful approach for studying protein-protein interactions in vivo. Here, we describe a protocol for antibody purification and immobilization followed by immunoprecipitation from plant tissue extracts using magnetic beads. The protocol has been used to detect regulators in the Zea mays phenylpropanoid pathway. The protocol is amenable to a variety of downstream assays, including western blotting and mass spectrometry.