荧光定量PCR检测基因枪轰击免疫小鹅瘟病毒VP3基因疫苗在小鼠体内的动态分布  

Fluorescent Quantitative PCR Detects the Dynamic Distribution of GPV-VP3 Gene Vaccine in Mice Vaccinated via Gene Gun Bombing

作　　者：刘晓东[1] 程安春[1] 汪铭书[1] 韩新锋[1] 卢菲[1] 黎敏[1] 陈孝跃[1] 

机构地区：[1]四川农业大学动物医学院禽病防治研究中心

出　　处：《四川农业大学学报》2007年第4期457-461,474,共6页Journal of Sichuan Agricultural University

基　　金：国家科技攻关重大项目(2004BA901A03);教育部"新世纪优秀人才支持计划"项目(NCET-04-0906);四川省重大基础研究项目(05JY029-109);四川省重点建设学科项目(SZD0418)

摘　　要：开展了检测小鹅瘟病毒(GPV)VP3基因的荧光定量PCR(FQ-PCR)的建立和基因枪轰击不同剂量(6μg/只、3μg/只和1μg/只)GPV-VP3基因疫苗(pcDNA-GPV-VP3)在BALB/c小鼠各组织器官(心、肝、脾、肺、肾、脑、肠和免疫部位皮肤)分布规律的研究。结果表明:①建立的FQ-PCR特异性强、灵敏度高、重复性好,核酸模板数与FQ-PCR测定的Ct值具有很好的直线相关性(相关系数达到0.999);②pcDNA-GPV-VP3在各剂量免疫小鼠1h即可在各组织中被检测到,其中在免疫部位皮肤含量最高,在心与肺中含量也较高,在脑中含量最低;③pcDNA-GPV-VP3在组织器官里的含量于3h开始下降,31wk仍能在3个剂量免疫组小鼠的各个组织器官中检测到,但多数组织器官中的含量比1h时约少了102,免疫部位皮肤减少了103;④不同剂量免疫组各组织器官中pcDNA-GPV-VP3含量6μg/只组>3μg/只组>1μg/只组,但剂量组之间的差异并不显著(P>0.05)。研究表明:FQ-PCR是定量检测pcDNA-GPV-VP3在免疫小鼠各组织器官含量的可靠实验手段,pcDNA-GPV-VP3免疫小鼠后1h时可分布至小鼠体内各组织器官中并持续存在31wk以上。In this paper we established the method of Fluorescent quantitative PCR （FQ- PCR） for detecting VP3 gene of Gosling plague virus （GPV） and studied the dynamic distribution of GPV- VP3 gene vaccine （pcDNA- GPV- VP3） in BALB/c mice tissues （cardiac muscle, liver, spleen, lung, kidney, brain, intestine and cutis of immune site） vaccinated via gene gun bombing with different dose （6μg per mouse, 3μg per mouse and 1μg per mouse）. The results showed that： ①The method of FQ - PCR is specific, sensitive and good repeated. There was a good linear correlation between the copy numbers of the nucleic acid and Ct value detected by FQ-PCR （correlation coefficients= 0. 999）; ② pcDNA- GPV - VP3 was detected in all tissues 1 h post-inoculation. The copy numbers of pcDNA- GPV-VP3 was largest in cutis of immune site, and very high in cardiac muscle and lung, and the least in brain; ③The copy numbers of pcDNA - GPV- VP3 in tissues began to decrease 3 h post-inoculation, pcDNA- GPV - VP3 was still detected in all tissues 31 wk post-inoculation, but the copy numbers decreased about 1.02 in most tissues and about 103 in cutis of immune site than ｜ h tissues; ④ The ranking of the copy numbers of pcDNA-GPV- VP3 in the same tissues was 6μg per mouse〉3 μg per mouse〉1μg per mouse, but the difference among them was not significant （P 〉0.05）. The research demonstrated that FQ - PCR was a reliable experiment tool for detecting the copy numbers of pcDNA-GPV- VP3 in mice, and pcDNA - GPV - VP3 could distribute to all over mice bodies lh and existed more than 31 wk post-inoculation.

关 键 词：荧光定量PCR 小鹅瘟病毒VP3基因疫苗 基因枪 小鼠 动态分布 

分 类 号：S858.335[农业科学—临床兽医学]