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

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。

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

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(6 μg per mouse,3 μg per mouse and 1 μg 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);②pcDNA-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 102 in most tissues and about 103 in cutis of immune site than 1 h tissues;④The ranking of the copy numbers of pcDNA-GPV-VP3 in the same tissues was 6 μg per mouse>3 μg per mouse>1 μg 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 1h and existed more than 31 wk post-inoculation.