Amplification and cloning of arabidopsis 6xhis-tagged mpk6 fusion encoded gene to characterize biochemical mitogen-activated protein kinase in disease resistance role against Fusarium graminearum

Abstract

The Mitogen-Activated Protein Kinase (MPK) cascade plays an important role in the intracellular signaling transduction pathway leading to resistance against phytopathogens produced by Fusarium graminearum. In the cascade, there are three prominent kinase protein groups involved, an MPK kinase kinase (MPKKK), MPK kinase (MPKK), and an MPK. Recognitions of pathogen-derived molecules in plants trigger rapid activation of some MPKs including MPK6 which are found in a wide variety of plant species, including in Arabidopsis thaliana. The structure of MPK6 contains kinase domain and common docking (CD) domain. CD domain is phosphorylated by interact with MPKK. Moreover, the MPKK which binds to MPK6 and its phosphorylation mechanism are still unknown, so as initial study is needed to investigate biochemical characterization by prepare MPK6 protein. In this research, mpk 6 was amplified by using a pair primer and subsequently was ligated into pET160/GW/D-TOPO vector which contained with sequence encoded 6xHistidine tag protein for protein purification assay.