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首页> 外文期刊>Virology >Characterization of recombination events leading to the production of an ecotropic replication-competent retrovirus in a GP+envAM12-derived producer cell line.
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Characterization of recombination events leading to the production of an ecotropic replication-competent retrovirus in a GP+envAM12-derived producer cell line.

机译：重组事件的特征导致在GP + envAM12衍生的生产细胞系中产生具有亲复制能力的逆转录病毒。

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摘要

Replication-competent retrovirus (RCR) was identified in a GP+envAM12-derived producer cell, containing the MFG-S-Neo retroviral vector, using a marker rescue assay. Studies were undertaken to determine the origin and structure of this RCR. Receptor interference assays demonstrated that the virus was pseudotyped with an ecotropic envelope. Molecular analysis demonstrated the presence of a MoMLV ecotropic env recombinant where the neomycin resistance gene of the MFG-S-Neo vector was replaced by MoMLV ecotropic env. Additional recombinants linking the retroviral pol gene to neo and the neo gene to MoMLV env were also identified. A full-length MoMLV retroviral genome was detected by nested PCR in the contaminated amphotropic producer cells and in cells infected with its supernatant. Unexpectedly, this was also present in the GP+E86 packaging cells together with a previously undescribed envelope construct possessing a full 5' and 3' LTR, although these cells were consistently negative for the presence of RCR. These anomalies in the GP+E86 packaging cell line result in increased homology with the MFG-S-Neo vector, leading to an increased risk for the production of RCR. Our findings point to a need for increased vigilance when using these packaging lines to generate replication-defective retrovirus. Copyright 2000 Academic Press.
机译：使用标记物拯救试验，在包含MFG-S-Neo逆转录病毒载体的GP + envAM12衍生的生产细胞中鉴定了具有复制能力的逆转录病毒（RCR）。进行了研究以确定该RCR的起源和结构。受体干扰分析表明，该病毒是带有亲热膜的假型。分子分析表明存在MoMLV亲环境env重组体，其中MFG-S-Neo载体的新霉素抗性基因被MoMLV亲环境env替代。还鉴定了将逆转录病毒pol基因与neo连接和将neo基因与MoMLV env连接的其他重组体。通过巢式PCR在受污染的两性生产细胞和上清液感染的细胞中检测到了全长MoMLV逆转录病毒基因组。出乎意料的是，这也存在于GP + E86包装细胞中，连同先前未描述的具有完整5'和3'LTR的包膜构建物，尽管这些细胞对于RCR的存在始终是阴性的。 GP + E86包装细胞系中的这些异常导致与MFG-S-Neo载体的同源性增加，从而导致产生RCR的风险增加。我们的发现表明，在使用这些包装线生成复制缺陷型逆转录病毒时，需要提高警惕。版权所有2000学术出版社。

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