人工合成根特异启动子SRSP的功能分析

Construction and function of a root-specific promoter SRSP

摘要:

根负责吸收水分和养分，是重要的植物组织，但易受生物及非生物胁迫，影响作物的生长发育和产量。设计合成根特异启动子，可为与胁迫相关的抗性基因在作物根部的功能研究及高效表达提供候选启动子。文中将4拷贝的根特异性顺式作用元件 (OSE1ROOTNODULE、OSE2ROOTNODULE、SP8BFIBSP8AIB和ROOTMOTIFAPOX1) 以串联排列方式设计合成了一个根特异性模块 (pro-SRS)，并与来自CaMV 35S启动子的最小启动子融合，合成一个人工合成启动子SRSP。通过替换CaMV 35S启动子将SRSP启动子克隆到pCAMBIA2300.1中以驱动GUS表达。将携带SRSP启动子的构建体通过农杆菌介导的方法转化到烟草中。GUS组织化学染色分析和实时PCR (RT-PCR) 分析显示SRSP启动子在转基因烟草中赋予根特异性表达。说明顺式作用元件重复排列可实现启动子预期功能，本研究为理性设计植物组织特异启动子奠定了理论基础。

Abstract:

The responsibility of root is absorbing water and nutrients, it is an important plant tissue, but easily to be affected by biotic and abiotic stresses, affecting crop growth and yield. The design of a synthetic root-specific promoter provides candidate promoters for the functional analysis and efficient expression of stress-related genes in crop roots. In this study, a synthetic root-specific module (pro-SRS) was designed using tandem four-copies of root specific cis-acting elements (OSE1ROOTNODULE, OSE2ROOTNODULE, SP8BFIBSP8AIB, and ROOTMOTIFAPOX1), and fused with minimal promoter from the CaMV 35S promoter to synthesize an artificially synthetic SRSP promoter. The SRSP promoter was cloned in pCAMBIA2300.1 by replacing CaMV 35S promoter so as to drive GUS expression. The constructs with SRSP promoter were transformed in tobacco by Agrobacterium-mediated method. SRSP promoter conferred root-specific expression in transgenic tobacco plants through Real-time PCR (RT-PCR) analysis and GUS histochemical staining analysis. It is indicated that the repeated arrangement of cis-acting elements can realize the expected function of the promoter. This study laid a theoretical foundation for the rational design of tissue-specific promoters.