Development of a Method for Visualizing and Quantifying ofEndogenous RNAs by Bioluminescent Probes

摘要

Gene expression plays important roles in physiological and pathophysiological conditions.Phenotypic changes such as tumorigenesis,differentiation,and reprogramming are caused bychanges in the expression of particular genes upon stimulation.For comprehension of themechanisms of the phenotypic changes,analysis of gene expression alteration is important.However,it is difficult to quantify temporal changes of specific gene expression in livingsamples at single cells.In this study,we aim to develop a method for temporal monitoring of aspecific RNA in single cells by luminescence observation.We designed a luminescence-basedRNA probe which consists of two fusion proteins with each different domain;;One is an RNArecognition domain that binds to a specific RNA sequence,and the other is an split luciferasedomain that catalyzes the luminescent reaction of luciferin.The probe responds to the presenceof target RNAs by emiting a luminescence signal through complementation of split luciferasedomains (Fig).