减蛋综合征病毒全基因组DNA文库的构建及物理图谱分析

Molecular Cloning and Physical Mapping of Egg Drop Syndrome Virus

Ｅ Ｄ Ｓ Ｖ Ａ Ａ２ 株纯化 Ｄ Ｎ Ａ 经 Ｈｉｎｄ Ⅲ、 ＰｓｔⅠ和 Ｓｐｈ Ⅰ水解后， 分别提取 Ｄ Ｎ Ａ 片段并克隆至ｐ Ｂｌｕｅｓｃｒｉｐｔ Ｋ Ｓ（ ＋ ） 和ｐ Ｇ Ｅ Ｍ３ Ｚｆ （ ＋ ） 载体， 构建了 Ｅ Ｄ Ｓ Ｖ 全基因文库。根据 Ｅ Ｄ Ｓ Ｖ Ｄ Ｎ Ａ 片段和基因组 Ｄ Ｎ Ａ 电泳迁移率计算， Ｅ Ｄ Ｓ Ｖ 基因组大小为３２９ｋｂ ， 通过克隆片段酶谱鉴定， 杂交建立了 Ｅ Ｄ Ｓ Ｖ 限制性内切酶 Ｈｉｎｄ Ⅲ、 Ｐｓｔ Ⅰ和 Ｓｐｈ Ⅰ的物理图谱。

One local strain AA 2 of Egg Drop Syndrome isolated in China,was inoculated into duck embryoes and the viral DNA was extracted and purified.DNA fragments of the viral genome digested by restriction enzymes(Hind Ⅲ,Pst I and Sph Ⅰ),were cloned into the sites of pBluescript KS(+)and pGEM 3Zf(+)vectores,respectively.According to the sizes of the individual restriction enzyme fragments,the total size of the EDSV genome was calculated to be 32 9Kb.The physical maps were defined by analysis of the viral DNA fragments with endonucleases and hybridization using DNA fragment probes labelled with Digoxigenin dUTP.