弓形虫不同发育时期棒状体颈部蛋白5基因的表达研究

Study on Expression of Toxoplasma gondii Rhoptry Neck Protein 5 Gene in Different Developmental Stages

为探究不同发育时期弓形虫棒状体颈部蛋白5(Toxoplasma gondii rhoptry neck protein 5,TgRON5)基因的表达情况与其毒力的相关性,本试验以Ⅱ型弓形虫PRU虫株作为研究对象,以弓形虫管家基因ACT1作为内参基因,分别对目的基因和内参基因设计特异性引物,通过对各基因建立标准曲线,确定二者扩增效率的一致性后,采用实时荧光定量PCR相对定量法对弓形虫4个不同发育时期(速殖子、缓殖子、未孢子化卵囊及孢子化卵囊)中TgRON5基因的表达情况进行检测与分析。结果显示,TgRON5基因在弓形虫各发育时期均有表达,其中,孢子化卵囊表达水平最高,未孢子化卵囊次之,速殖子较低,缓殖子最低,表明RON5蛋白的合成与分泌与虫体入侵宿主细胞的侵袭力有着密切的关系。本研究结果为阐明TgRON5蛋白参与弓形虫入侵的机理奠定了基础。

To investigate the relationship between the expression level of Toxoplasma gondii rhoptry neck protein 5（TgRON5）gene in different developmental stages and the virulence of Toxoplasma gondii,the objective of this study was to examine the different expression of TgRON5 gene in different developmental stages of typeⅡ Toxoplasma gondii PRU strain.Specific primers were designed according to the sequence of TgRON5 gene,and ACT1 gene of Toxoplasma gondii was used as a reference gene.Following the establishment of standard curve for the target and reference genes,in order to confirm the consistency of their amplification efficiency,Real-time PCR method was applied to determine and compare the expression level of TgRON5 gene in development stage which including tachyzoite,bradyzoite,non-sporulated oocyst and sporulated oocyst.The results demonstrated that TgRON5 was expressed in all develop-mental stages but the sporulated oocysts had the highest expression,followed by the non-sporulated oocysts,the third was tachyzoite,and the lowest was bradyzoite.It suggests that the synthesis and secretion of TgRON5 protein was closely associated with the invasiveness of the parasite.All these findings had important implications for elucidating the functions of TgRON5 involved in the invasion of Toxoplasma gondii.