Optimized phos-tag mobility shift assay for the detection of protein phosphorylation in planta

• 발행기관: (사)한국식물생명공학회
• 저자명: Shah Hussain Nhan Thi Nguyen Xuan Canh Nguyen Chae Oh Lim Woo Sik Chung
• 간행물 정보: 『Journal of Plant Biotechnology』45권 4호, 322~327쪽, 전체 6쪽
• 주제분류: 농수해양 > 기타농수해양
• 파일형태: PDF
• 발행일자: 2018.12.31

영문 초록

Post-translational modification of proteins regulates signaling cascades in eukaryotic system, including plants. Among these modifications, phosphorylation plays an important role in modulating the functional properties of proteins. Plants perceive environmental cues that directly affect the phosphorylation status of many target proteins. To determine the effect of environmentally induced phosphorylation in plants, in vivo methods must be developed. Various in vitro methods are available but, unlike in animals, there is no optimized methodology for detecting protein phosphorylation in planta. Therefore, in this study, a robust, and easy to handle Phos-Tag Mobility Shift Assay (PTMSA) is developed for the in vivo detection of protein phosphorylation in plants by empirical optimization of methods previously developed for animals. Initially, the detection of the phosphorylation status of target proteins using protocols directly adapted from animals failed. Therefore, we optimized the steps in the protocol, from protein migration to the transfer of proteins to PVDF membrane. Supplementing the electrophoresis running buffer with 5 mM NaHSO3 solved most of the problems in protein migration and transfer. The optimization of a fast and robust protocol that efficiently detects the phosphorylation status of plant proteins was successful. This protocol will be a valuable tool for plant scientists interested in the study of protein phosphorylation.

목차

Introduction

Materials and Methods

Results and Discussion

Acknowledgments

Reference

키워드

Electrophoresis mobility shift assay Phos-tag Plant Protein kinases Protein phosphorylation