Recombinant Epstein-Barr virus with small RNA (EBER) genes deleted transforms lymphocytes and replicates in vitro.
Recombinant Epstein-Barr virus with small RNA (EBER) genes deleted transforms lymphocytes and replicates in vitro.
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DOI：
10.1073/pnas.88.4.1546
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发表时间：
1991-02
影响因子：
11.1
通讯作者：
S. Swaminathan;B. Tomkinson;E. Kieff
中科院分区：
文献类型：
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作者：
S. Swaminathan;B. Tomkinson;E. Kieff
使用 EBV P3HR-1 菌株通过同源重组制备了删除了小 RNA (EBER) 基因的 Epstein-Barr 病毒 (EBV) 菌株，该菌株已删除了编码 EBV 核抗原 EBNA-2 的必需转化基因，以及 EBNA-2 基因座处为野生型但 EBER 基因已被删除的 DNA 片段。尽管 EBER 和 EBNA-2 基因相隔 40 KB，但选择需要恢复 EBNA-2 基因的转化 P3HR-1 重组体会导致 20% 的 EBER 缺失共转移。 EBER 缺失的重组体将原代 B 淋巴细胞转化为类淋巴母细胞系 (LCL)，这些细胞与野生型 EBV 转化的 LCL 在增殖、潜伏期相关 EBV 基因表达以及对 EBV 复制周期基因表达的允许性方面没有区别。来自受感染 LCL 克隆的 EBER 缺失病毒可以感染原代 B 淋巴细胞并使其生长转化。这些程序应适用于 EBNA-2 基因 40 KB 以内的其他 EBV 重组体的构建。 EBER 缺失的 EBV 重组体应该有助于进一步评估 EBER 在 EBV 感染中的作用。
Strains of Epstein-Barr virus (EBV) with deletions of the small RNA (EBER) genes were made by homologous recombination using the EBV P3HR-1 strain, which has undergone deletion of the essential transforming gene that encodes the EBV nuclear antigen, EBNA-2, and a DNA fragment that was wild type at the EBNA-2 locus but from which the EBER genes had been deleted. Even though the EBER and EBNA-2 genes are separated by 40 kilobases, selection for transforming P3HR-1 recombinants that required a restored EBNA-2 gene resulted in 20% cotransfer of the EBER deletion. EBER-deleted recombinants transformed primary B lymphocytes into lymphoblastoid cell lines (LCLs), which were indistinguishable form LCLs transformed by wild-type EBV in their proliferation, in latency-associated EBV gene expression, and in their permissiveness for EBV replication cycle gene expression. EBER-deleted virus from infected LCL clones could infect and growth-transform primary B lymphocytes. These procedures should be applicable to the construction of other EBV recombinants within 40 kilobases of the EBNA-2 gene. The EBER-deleted EBV recombinants should be useful in further evaluating the role of EBERs in EBV infection.