鸡传染性支气管炎病毒S1基因在昆虫细胞中表达水平初探

将含有鸡传染性支气管炎病毒 Ｓ１ 基因ｃ Ｄ Ｎ Ａ 的重组转移质粒ｐ Ｓ Ｘ Ｉ Ｖ Ｖ Ｉ＋ Ｘ３ Ｓ１ ． Ｈｏｌｔｅ 和ｐ Ｓ Ｘ Ｉ Ｖ Ｖ Ｉ＋ Ｘ３／４ Ｓ１ ． Ｈｏｌｔｅ 分别与粉纹夜蛾核型多角体病毒 Ｔｎ Ｎ Ｐ Ｖ Ｓ Ｖ Ｉ－ Ｇ Ｄ Ｎ Ａ（ Ｏ Ｃ Ｃ－ ，ｇａｌ＋ ） 共转染草地夜蛾（ Ｓｆ９） 细胞，经空斑纯化得到重组病毒 Ｔｎ Ｎ Ｐ Ｖ（ Ｘ３） Ｓ１ ． Ｈｏｌｔｅ Ｏ Ｃ Ｃ＋ 和 Ｔｎ Ｎ Ｐ Ｖ（ Ｘ３／４） Ｓ１ ． Ｈｏｌｔｅ Ｏ Ｃ Ｃ＋ 。将重组毒株分别感染 Ｔｎ５ Ｂ１ 细胞，并进行 Ｓ Ｄ Ｓ Ｐ Ａ Ｇ Ｅ 与 Ｗｅｓｔｅｒｎｂｌｏｔ 检测。结果表明， Ｔｎ Ｎ Ｐ Ｖ（ Ｘ３／４） Ｓ１ ． Ｈｏｌｔｅ Ｏ Ｃ Ｃ＋ 在感染的细胞中高效表达了 Ｓ１ 蛋白， Ｓ Ｄ Ｓ Ｐ Ａ Ｇ Ｅ 凝胶薄层色谱分析结果显示，感染病毒后７２ ｈ Ｓ１ 蛋白的表达量占细胞内总蛋白量的３５ ．８ ％ ，而 Ｔｎ Ｎ Ｐ Ｖ（ Ｘ３） Ｓ１ ． Ｈｏｌｔｅ Ｏ Ｃ Ｃ＋ 感染的细胞内检测不出 Ｓ１ 蛋白。经分析认为这一差异主要来自 Ｓ１ 基因翻译起始位点及其附近的周围环境。

Preliminary Study on Expression Level of the S1 Gene of Avian Infectious Bronchitis Virus in a Baculovirus System

The recombinant transfer plasmids pSXIVVI\ X3 S1.Holte and pSXIVVI\ X3/4 S1.Holte containing S1 gene cDNA of avian infectious bronchitis virus and the parent virus (TnNPV SVI -G) DNA were used to co transfect \%Spodotera frugiperda\% (Sf) cells. The recombinant viruses TnNPV (X3)S1.Holte OCC and TnNPV (X3/4)S1.Holte OCC , which included the S1 gene and could form polyhedra, were plaque purified. The expression products of S1 gene were detected by SDS PAGE and Western blotting in Tn 5B1 cells infected with the recombinant viruses. The S1 protein with a molecular weight of 100 kD was expressed in the cells infected with TnNPV (X3/4)S1.Holte OCC at high efficiency, the expression level in Tn 5B1 cells 72 h postinfection being 35.8% by SDS PAGE gel thin layer chromatography. No expression product was detected by SDS PAGE and Western blotting in Tn 5B1 cells infected with TnNPV (X3)S1.Holte OCC . The expression level depends mainly upon the translational start site of the S1 gene and its flanking sequences.