Allison R F, Janda M, Ahlquist P
Institute of Molecular Virology, University of Wisconsin-Madison 53706.
J Virol. 1988 Oct;62(10):3581-8. doi: 10.1128/JVI.62.10.3581-3588.1988.
Complete cDNA copies of genomic RNA1, RNA2, and RNA3 of cowpea chlorotic mottle virus (CCMV) were cloned 1 base downstream from a T7 RNA polymerase promoter. The mixture of capped in vitro transcripts from all three clones produced normal CCMV infections in barley protoplasts and cowpea plants. By using transcripts from these clones and from a similar set of biologically active clones of the related brome mosaic virus (BMV), all possible single component exchanges between the BMV and CCMV tripartite genomes were tested. Viral RNA replication was not observed with any heterologous combination of RNA1 and RNA2, which encode trans-acting viral RNA replication factors. However, substitution of the heterologous RNA3 into either genome produced viable hybrid viruses, both of which replicated in barley protoplasts and produced lesions on Chenopodium hybridum, a local lesion host for both parent viruses. In hybrid infections, BMV and CCMV coat proteins each readily packaged RNAs from the heterologous virus, but BMV RNAs were replicated to a higher level than CCMV RNAs, even when trans-acting RNA replication factors were provided by CCMV genes. Neither hybrid systemically infected the natural host of either parent virus, suggesting that host specificity determinants in BMV and CCMV are encoded by RNA3 and at least one other genomic RNA.
将豇豆褪绿斑驳病毒（CCMV）的基因组RNA1、RNA2和RNA3的完整cDNA拷贝克隆到T7 RNA聚合酶启动子下游1个碱基处。来自所有三个克隆的加帽体外转录本混合物在大麦原生质体和豇豆植株中产生了正常的CCMV感染。通过使用这些克隆以及相关的雀麦花叶病毒（BMV）一组类似的具有生物活性的克隆的转录本，对BMV和CCMV三方基因组之间所有可能的单组分交换进行了测试。对于编码反式作用病毒RNA复制因子的RNA1和RNA2的任何异源组合，均未观察到病毒RNA复制。然而，将异源RNA3替换到任一基因组中产生了有活力的杂交病毒，这两种杂交病毒都在大麦原生质体中复制，并在双亲病毒的局部病斑寄主杂种藜上产生病斑。在混合感染中，BMV和CCMV外壳蛋白都很容易包装来自异源病毒的RNA，但即使CCMV基因提供反式作用RNA复制因子，BMV RNA的复制水平也高于CCMV RNA。这两种杂交病毒均未系统感染任一亲本病毒的天然寄主，这表明BMV和CCMV中的寄主特异性决定因素由RNA3和至少一种其他基因组RNA编码。
