摘要
在已有的全长感染性克隆pFD3的基础上,构建了新的低拷贝的全长克隆pLGFD3-8。按照疫苗制备过程中env基因的变化情况,采用基因替换和定点突变的方法,构建了一系列具有马传染性贫血病毒(EIAV)强毒株env基因及其主要突变特征的嵌合克隆。利用这些克隆转染FDD细胞,并用逆转录酶活性检测和PCR方法确定其感染性。结果发现,在FDD细胞中传代3次后,可在细胞培养物中检测到逆转录酶活性和原病毒DNA的存在,在电镜下可以观察到典型的EIAV病毒颗粒。这一结果为进一步研究马传染性贫血病毒致病的分子机制和免疫保护机理奠定了良好的基础。
On the basis of the infectious clone pFD3,we constructed another full-length clone pLGFD3 with low copy numberAccording to the variation of env gene during vaccine preparation,several chimeric clones with env gene or with main characteristics of EIAV wild strains were constructed by gene substitution and site-directed mutagenesis methodsThese clones were used to transfect fetal donkey dermal(FDD) cell and then their infectious characteristics were monitored by PCR test and RT activity assayAfter 3 generations passaged in FDD cell,the RT activity and the provirus DNA could be detected in cell cultureThe EIAV particles could be observed under electron microscope in cell cultureAll of these provided a solid basis for further study of the pathogenic mechanisms of EIAV and the immune protection of Chinese EIAV vaccines
出处
《病毒学报》
CAS
CSCD
北大核心
2003年第2期128-132,共5页
Chinese Journal of Virology
基金
基础研究重大项目前期研究专项(2001CCA00600)
国家攀登计划特别项目
关键词
感染性马传染性贫血病毒
嵌合克隆
基因替换
定点突变
感染性克隆
equine infectious anemia virus(EIAV)
gene substitution
site-directed mutagenesis
env gene
infectious clone
