摘要
为构建表达鸡IL-18的重组禽痘病毒,将含痘病毒启动子LP2EP2驱动的鸡IL-18基因插入到禽痘病毒转移载体pSY681中,获得重组转移载体pSY681/ChIL-18。将pSY681/ChIL-18转染已感染亲本禽痘病毒S-FPV-017株的鸡胚成纤维细胞,使其在鸡胚成纤维细胞内与禽痘病毒基因组发生同源重组,产生表达鸡IL-18的重组禽痘病毒rFPV-ChIL-18。在含有X-gal的营养琼脂培养基上进行蓝斑筛选后,对重组病毒rFPV-ChIL-18又进行了多次蚀斑克隆。以重组禽痘病毒DNA为模板,利用鸡IL-18基因特异引物进行PCR,扩增出1条约0.6 kb的带。收集含鸡IL-18蛋白的细胞上清液进行MTT试验,表达的产物能明显提高鸡淋巴细胞的转化率。
In order to construct the recombinant fowlpox viruses（rFPV） expressing chicken IL-18, the chicken IL-18 cDNA was cloned into EcoR Ⅰ site of pSY538 plasmid and then subcloned into Sfi Ⅰ site of pSY681 plasmid containing LacZ gene to generate the recombinant plasmid pSY681/ChIL-18. The recombinant plasmid was transfected on the chicken embryo fibroblasts cell（ CEF）that was pre-infected with S-FPV-017 .By screening of blue plaques on the CEF overlaid with agar containing X-gal,the rFPV-ChIL-18 recombinants were obtained and identified by PCR. Recombinant chicken IL-18 induced in vitro the proliferation of ConA-stimulated chicken splenocytes in MTT assay.
出处
《华北农学报》
CSCD
北大核心
2008年第6期6-10,共5页
Acta Agriculturae Boreali-Sinica
基金
国家“十一五”科技支撑计划专项(2006BAD06A08)
关键词
鸡IL-18基因
重组禽痘病毒
构建
生物学活性
Chicken interleukin-18 gene
Recombinant fowlpox virus
Construction
Biologic activity
