神经营养素-3重组腺病毒Ad-NT-3的培养和鉴定

目的 培养和鉴定重组腺病毒载体Ａｄ－ＮＴ－３。方法 在２９３细胞中培养扩增Ａｄ－ＮＴ－３,用组织培养半数感染量测测定其滴度。从病毒培养上清中提取ＤＮＡ,用聚合酶链反应技术扩增ＮＴ－３部分基因片段,将扩增产物克隆并进行序列分析以测定Ａｄ－ＮＴ－３中ＮＴ－３的存在及种属。结果 Ａｄ－ＮＴ－３扩增后获得了较高滴度的病毒。从形态学证实Ａｄ－ＮＴ－３扩增后获得了较高滴度的病毒。从形态学证实Ａｄ－ＮＴ－３含有

Propagation and identification of adenovirus vector for neurotrophin-3

AIM To propagate and to identify neurotrophin-3 recombinant adenovirus vector. METHODS Recombinant adnoviral vector for NT-3 was propagated in 293 packaging cells. Viral titers were determined by tissue culture infections dose50 (TCID_ 50) method. Polymerase chain reaction (PCR) and sequence analysis were employed to confirm the presence of the NT-3 cDNA in the genome of the adenovirus vector and its species. RESULTS High titers of recombinant adenoviral particles after propagation of Ad-NT-3 in 293 cells were obtained. The presence of replication competent adenovirus was determined morphologically by infecting 293 cells with different dilutions of Ad-NT-3 and monitoring them during 10 days for appearance of a cytopathic effect (CPE). Ad-NT-3 contains the gene of NT-3, which was human NT-3 by origin. CONCLUSION There is human NT-3 gene and replication competent adenovirus in Ad-NT-3.