斑节对虾白斑综合症病毒部分基因组文库及核酸探针检测法
Partial cloning of the genome library of white spot syndrome virus from Penaeus monodon and DNA probe detection methods
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                            摘要: 通过分离纯化白斑综合症病毒( WSSV) 粒子, 抽提病毒DNA, 用限制性内切酶EcoR. 或Sal .酶切后, 克隆入质粒pBluescript òKS 中, 从而建立了WSSV 部分基因组文库。估计WSSV 基因组DNA 在 165kb 以上。将WSSV EcoR. 克隆片段标记制备为探针, 进行Southern 杂交、打点杂交和原位杂交, 其结果证 明了克隆片段对WSSV 特异, 并为检测WSSV 提供了方法。通过对部分基因组文库序列分析发现,WSSV 核 酸序列与已知杆状病毒核酸序列的同源性低, 与GenBank 中已有病毒核酸序列比较都无较大同源性, 说明 WSSV 不属于杆状病毒科, 目前只能列为未分类的无脊椎动物病毒, 其分类地位尚需更多证据方能确定。Abstract: White spot syndrome virus(WSSV) , a non-occluded virus which is the causat ive agent of a disease caused high shrimp mortalities and severe damage to shrimp cultures, was iso lated and purified. Virions are rod-shaped, enveloped, 430nm @ 120nm in size, with a tai-l like appendage at one extremity. Nucleocapsids are 300nm @ 85nm in size, and are composed of rings of subunits in a stacked series. The genome of the virus is a double-stranded DNA molecule. The full length of the DNA is estimated to be longer than 165kb. EcoR . or Sal . restriction fragents obtained after agarose gel electrophoreses were cloned in the system pBluescript ò KS-XL1-blue. Southern hybridization, dot blot hybridization and in situ hybridizat ion were performed using the WSSV EcoR . clone fragments as probe, and those proved that these clone fragments are specific to WSSV. According to the results of nucleotide sequence analysis, none of the WSSV clones showed considerable se- quence homology with those of other known viruses or organisms, indicating that WSSV may only be distantly related to baculoviruses.
 
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