摘要
利用脂质体转染技术,将含有SNV株禽网状内皮组织增生症病毒(REV)前病毒全基因组cDNA克隆质粒转染鸡胚成纤维细胞(CEF)。用对REV的单克隆抗体和抗REV env-gp90的鼠血清作间接免疫荧光反应,在原始的转染细胞及随后传代的细胞中均显示病毒特异性抗原。而且,在连续传代细胞中的阳性率明显升高。用REV特异性引物对进一步传代后的细胞基因组作PCR,也检测出REV基因组。这些结果均表明所得到的分子克隆化病毒具有传染性,因而也进一步证明所用的质粒克隆包含有具感染性的全病毒基因组。对该全基因组cDNA克隆进行酶切所获得的数个亚克隆进行测序,并将序列进行拼接,完成了REV全基因组序列。REV的这个传染性克隆将有助于进一步研究REV的分子生物学特性。
Chicken embryo fibroblast cells （CEF） were transfected with avian reticuloendotheliosis virus （REV） proviral eDNA clone mixed with lipofectamine. In indirect immune fluorescence antibody test with monoclonal antibody specific to REV or with mouse antiserum against REV env gp90 protein expressed in E. coli , REV antigen was detected in both primary transfected CEF and continually passaged cells inoculated with culture supernatants of primary transfected CEF cells by IFA. By PCR, the REV gp90 gene could also be amplified from each passage of CEF inoculated with cell culture supernatants. The results indicated that the recombinant plasmid containing intact REV genome eDNA was able to produce infectious virus in CEF. The SNV proviral eDNA clone plasmid was sequenced, the infectious clone and its complete sequence of SNV strain would be useful for further study of REV at the molecular level.
出处
《病毒学报》
CAS
CSCD
北大核心
2005年第6期448-455,共8页
Chinese Journal of Virology
基金
国家自然科学基金重点项目(30330450)
