摘要
利用重组表达质粒ＨＮ－ＰＥＦＬ－２９与鸡痘病毒ＦＰ９共转染鸡胚成纤维细胞，经蚀斑筛选，结合Ｓｏｕｔｈｅｒｎ杂交，获得含有外源ＨＮ基因的纯化重组病毒。将筛选到的重组病毒经４次传代，细胞可见明显病变。免疫荧光检测结合Ｗｅｓｔｅｒｎ印迹表明该重组子表达产物主要位于细胞膜上，以糖基化（７４ｋＤ）和非糖基化（６８ｋＤ）２种形式存在，表达产物具有类似于天然ＮＤＶＨＮ蛋白的红细胞吸附活性及神经氨酸酶活性，且这２种活性均可被ＮＤＶＨＮ特异性单抗所抑制。对该表达产物的免疫活性研究表明：用重组病毒免疫４周龄ＳＰＦ鸡的ＥＬＩＳＡ抗体水平和血凝抑制效价均高于ＦＰＶ接种组，但低于ＬａＳｏｔａ弱毒活疫苗接种组。以ＮＤＶ强毒株Ｆ４８Ｅ９攻毒发现：接种重组病毒１０６ｐｆｕ／只时，保护力可达８０％，而１０４ｐｆｕ／只时，保护率仅为６０％，这表明该重组病毒对雏鸡具有一定的保护作用，且该保护作用与接种剂量有一定的相关性。
recombinant fowlpox virus transfer vector was constructed by cloning HaemagglutininNeuraminidase(HN) cDNA gene of Newcastle disease virus strain F48E9 under control of P7.5 promotor. The vector was contransfected into chicken embryo cells with FP9. Postive recombinants were selected by three cycles of plaque pruification and Southern blot. By immunofluorescence and Western blot, chicken embryo cells transfected with recombinants expressed two kinds of HNsized proteins(MW 74 kD; MW 68 kD).Both of them were HNspecific proteins and could react with antiHN antibody. Their molecular weight were conincided with glycosylated and unglycosylated NDV HN proteins. Cells transfected with recombinanat rFPVHN could adsorb chicken red blood cells and the cells surfaces exhibited neuraminidase activity. Both of these activities can be inhibited by antiHN antibody. Four groups of SPF chickens were inoculated with the cells infected with the recombinant virus, La Sota and FP9 respectively. Chickens inoculated with the recombinant virus developed antibodies against NDV. Though the antibody titers were not as high as that of chickens inoculated with NDV La Sota, they can protect chickens against the NDV F48E9 challenge.
出处
《中国免疫学杂志》
CAS
CSCD
北大核心
1998年第1期61-64,共4页
Chinese Journal of Immunology
基金
国家"八五"攀登计划
关键词
新城疫病毒
HN基因
重组禽痘病毒
Newcastle disease virus
HN gene
Recombinant fowlpox virus
