Construction of Toxoplasma ROP18 recombinant adenovirus vector and its effect on the apoptosis of C17.2 stem cells
ZHANG Xian1, 2, GAN Xiao-feng1, 2, CHENG Zheng-yang1, 2, DU Jian3, LUO Qing-li1, 2, SHEN Ji-long1, 2, YU Li1, 2
1.Department of Microbiology and Parasitology, Auhui Medical University, Hefei 230032,China; 2.Anhui Key Laboratory of Zoonoses, Auhui Medical University, Hefei 230032, China; 3.Department of Biochemistry, Auhui Medical University, Hefei 230032, China
Abstract：To study the effect of Toxoplasma rhoptry kinase 18 (ROP18) on the apoptosis of C17.2 neural stem cells, Toxoplasma gondii ROP18 was subcloned from PEGFP-G2-ROP18 into pHBAd-MCMV-GFP. The recombinant adenovirus vector pHBAd-MCMV-GFP-ROP18 was comfirmed by PCR and sequencing, and the identified recombinant adenovirus vector was transfected into 293 cells for packaging with skeleton plasmid. After amplification, ROP18 recombinant adenovirus (MOI=70) was transfected into C17.2 neural stem cell. The proliferation of C17.2 cells at 48 horus Post-treatment was detected using CCK-8 kit and the apoptosis of C17.2 cells at 24 hours Post-treatment was detected by flow cytometer. Western blot was used to determine the expression levels of caspase-3. ROP18 recombinant adenovirus was expressed effectively in C17.2 neural stem. Compared with the control group, proliferation of C17.2 cells was significantly decreased (P<0.05) in the ROP18 recombinant adenovirus group at 48 hours while the apoptosis of C17.2 cells was significantly increased in the ROP18 recombinant adenovirus (P<0.001) at 24 hours Post treatment and the caspases-3 expression levels was significantly increased (P<0.05). ROP18 recombinant adenovirus was successfully constructed and was highly expressed in C17.2 neural stem cells. ROP18 recombinant adenovirus induced the apoptosis of C17.2 neural stem cells.
张贤, 甘小凤, 程正阳, 都建, 罗庆礼, 沈继龙, 余莉. 弓形虫ROP18重组腺病毒载体的构建及其对神经干细胞C17.2凋亡的影响[J]. 中国人兽共患病学报, 2016, 32(3): 234-239.
ZHANG Xian, GAN Xiao-feng, CHENG Zheng-yang, DU Jian, LUO Qing-li, SHEN Ji-long, YU Li. Construction of Toxoplasma ROP18 recombinant adenovirus vector and its effect on the apoptosis of C17.2 stem cells. Chinese Journal of Zoonoses, 2016, 32(3): 234-239.
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2016, Vol. 32 
