斜纹夜蛾NPV p74基因的克隆及部分序列分析

Cloning and Sequencing of p 74 Gene of Spodoptera litura Nuclear Polyhedrosis Virus

以含ｐ７４基因的ＡｃＮＰＶＥｃｏＲⅠ－Ｐ片段作探针，与ＳｌＮＰＶ基因组在非严格条件下进行Ｓｏｕｔｈｅｒｎ杂交，将ＳｌＮＰＶｐ７４基因定位于４９００ｂｐ的ＳｌＮＰＶＸｈｏⅠ－Ｐ片段．对ＳｌＮＰＶＸｈｏⅠ－Ｐ片段中的一段９４０ｂｐ的ＤＮＡ片段进行序列测定，通过ｉｎｔｅｒｎｅｔ经ＢＬＡＳＴ与Ｇｅｎ－Ｂａｎｋ中的ｄａｔａｂａｓｅ比较分析发现，序列中的一段２４３ｎｔ序列与ＡｃＮＰＶｐ７４基因有６０％的同源性，一段８１ｎｔ序列与ＡｃＮＰＶｐ７４基因的同源性高达７９％．与ＣｆＮＰＶｐ７４相比，２段２６４ｎｔ和１００ｎｔ序列分别有６０％和７１％的同源性．测得序列中的部分序列与ＢｍＮＰＶ、ＯｐＮＰＶｐ７４基因也有高达５８％～７８％的同源性．同时，这部分序列编码的氨基酸与ＡｃＮＰＶ及ＯｐＮＰＶＰ７４蛋白的氨基酸序列也有很高的同源性．结果表明所测序列的一部分为ＳｌＮＰＶＰ７４蛋白Ｃ－端的编码序列．

Based on Southern blot hybridization at low stringency using the Autographa califonica nuclear polyhedrosis virus( Ac NPV) p 74 gene as heterologous probe( Eco RⅠ fragment P of Ac NPV DNA containing partial p 74 gene), the p 74 gene of Spodoptera litura NPV( Sl NPV) were localized on the XhoⅠ fragment P (4 900 bp) of the physical map of the viral genome. A 938 nt fragment( Bam HⅠ Eco RⅠ) of Xho Ⅰ fragment P was subcloned and sequenced. Two fragments, 243 nt and 81 nt, from the determined sequence, shared 60% and 79% nucleotide identity with Ac NPV p 74 gene respectively. Another two fragments, 264 nt and 100 nt, shared 60% and 71% nucleotide identity with Choristoneura fumiferana NPV( Cf NPV) p 74 gene respectively. Partial sequence of the 938 nt fragment showed 58%￣78% nucleotide sequence homology to Bombyx mori NPV( Bm NPV) and Orgyia pseudotsugata NPV( Op NPV) p 74 genes. Amino acid sequence encoded by the partial determined nucleotide sequence shared high amino acid sequence similarity with Ac NPV and Op NPV P74 proteins. The results indicate that the partial sequence of the 938 nt fragment is the Sl NPV P74 protein coding sequence.