Microscale Thermophoresis Assay: A Powerful Method to Quantify Protein–Nucleic Acid and Protein–Protein Interactions

Plant viruses are among the most disastrous pathogens that cause enormous losses in crop yield and crop quality, threatening food security worldwide. These losses are attributed to virus-induced plant physiological disorders and diseases. During the ever-evolving arm race between viruses and their hosts, viruses have evolved multiple strategies to disrupt plant fortification, for example, taking advantage of RNA silencing and plant hormone pathways. Microscale thermophoresis (MST) assay arises as a rapid and precise method to identify the interactions between biomolecules including host and viral factors from plant virus infections.The technique is based on the motion of molecules in temperature gradients, which is termed thermophoresis, and depends on the binding-induced changes in molecular size, charge, and solvation shell. This chapter provides a detailed protocol describing the application of MST assays in analyzing protein-nucleic acid and protein-protein interactions, taking the Rice stripe virus (RSV )-encoded nonstructural protein 3 (NS3) that recognizes primary miR528 transcript (pri-miR528) and Rice dwarf virus (RDV )-encoded capsid protein P2 that recognizes OsIAA10 as examples.