Tahmasebi Amin-Alah, Afsharifar Alireza
Plant Virology Research Center, Shiraz University, Shiraz, Iran.
Mol Biol Res Commun. 2017 Jun;6(2):45-56.
Transient expression of proteins in plants has become a choice to facilitate recombinant protein production with its fast and easy application. On the other hand, host defensive mechanisms have been reported to reduce the efficiency of transient expression in plants. Hence, this study was designed to evaluate the effect of cap analog and Potato virus A helper component proteinase (PVA HC-Pro) on green fluorescent protein (GFP) expression efficiency. . leaves were inoculated with capped or un-capped RNA transcripts of a Turnip crinkle virus (TCV) construct containing a green fluorescent protein reporter gene (TCV-sGFP) in place of its coat protein (CP) ORF. PVA HC-Pro as a viral suppressor of RNA silencing was infiltrated in trans by , increased the GFP foci diameter to six and even more cells in both capped and un capped treatments. The expression level of GFP in inoculated plants with TCV-sGFP transcript pre-infiltrated with PVA HC-Pro was 12.97-fold higher than the GFP accumulation level in pre-infiltrated leaves with empty plasmid (EP) control. Also, the yield of GFP in inoculated plants with capped TCV-sGFP transcript pre-infiltrated with EP and PVA HC-Pro was 1.54 and 1.2-fold respectively, greater than the level of GFP expressed without cap analog application at 5 days post inoculation (dpi). In addition, the movement of TCV-sGFP was increased in some cells of inoculated leaves with capped transcripts. Results of this study indicated that PVA HC-Pro and mRNA capping can increase GFP expression and its cell to cell movement in .
蛋白质在植物中的瞬时表达因其快速简便的应用方式,已成为促进重组蛋白生产的一种选择。另一方面,据报道宿主防御机制会降低植物中瞬时表达的效率。因此,本研究旨在评估帽类似物和马铃薯A病毒辅助成分蛋白酶(PVA HC-Pro)对绿色荧光蛋白(GFP)表达效率的影响。用含有绿色荧光蛋白报告基因(TCV-sGFP)以取代其外壳蛋白(CP)开放阅读框的芜菁皱缩病毒(TCV)构建体的加帽或未加帽RNA转录本接种叶片。作为RNA沉默病毒抑制子的PVA HC-Pro通过……进行反式浸润,在加帽和未加帽处理中均将GFP病灶直径增加到六个甚至更多细胞。用PVA HC-Pro预浸润的TCV-sGFP转录本接种的植物中GFP的表达水平比用空质粒(EP)对照预浸润的叶片中GFP积累水平高12.97倍。此外,用EP和PVA HC-Pro预浸润的加帽TCV-sGFP转录本接种的植物中GFP的产量在接种后5天分别比未应用帽类似物时表达的GFP水平高1.54倍和1.2倍。另外,在有加帽转录本接种叶片的一些细胞中TCV-sGFP的移动增加。本研究结果表明,PVA HC-Pro和mRNA加帽可增加GFP在……中的表达及其细胞间移动。