Department of Plant Pathology, Irrigated Agriculture Research and Extension Center, Washington State University, WA 99350, United States.
Citrus Research and Education Center, University of Florida, Lake Alfred, FL, United States.
Virology. 2018 Oct;523:89-99. doi: 10.1016/j.virol.2018.07.023. Epub 2018 Aug 10.
Infectious cDNA clones were developed for Grapevine leafroll-associated virus 3 (GLRaV-3, genus Ampelovirus, family Closteroviridae). In vitro RNA transcripts generated from cDNA clones showed replication via the production of 3'-coterminal subgenomic (sg) mRNAs in Nicotiana benthamiana protoplasts. The detection of sgRNAs and the recovery of progeny recombinant virions from N. benthamiana leaves agroinfiltrated with full-length cDNA clones confirmed RNA replication and virion formation. The 5' non-translated region (5' NTR) of GLRaV-3 was exchangeable between genetic variants and complement the corresponding cognate RNA functions in trans. Mutational analysis of the 5' NTR in minireplicon cDNA clones showed that the conserved 40 nucleotides at the 5'-terminus were indispensable for replication, compared to downstream variable portion of the 5' NTR. Some of the functional mutations in the 5' NTR were tolerated in full-length cDNA clones and produced sgRNAs and virions in N. benthamiana leaves, whereas other mutations affected replication and virion formation.
葡萄卷叶伴随病毒 3（GLRaV-3，属马铃薯 Y 病毒科，杆状病毒科）的感染性 cDNA 克隆被开发出来。从 cDNA 克隆中生成的体外 RNA 转录本在本氏烟原生质体中通过产生 3'末端亚基因组（sg）mRNA 进行复制。在全长 cDNA 克隆 agroinfiltrated 的烟叶片中检测到 sgRNAs 和重组子代病毒粒子的回收，证实了 RNA 复制和病毒粒子形成。GLRaV-3 的 5'非翻译区（5'NTR）在遗传变异体之间可互换，并在转译中互补相应的同源 RNA 功能。对 minireplicon cDNA 克隆中 5'NTR 的突变分析表明，与 5'NTR 的下游可变部分相比，5'末端保守的 40 个核苷酸对于复制是必不可少的。5'NTR 中的一些功能突变在全长 cDNA 克隆中是可以容忍的，并在烟叶片中产生 sgRNAs 和病毒粒子，而其他突变则影响复制和病毒粒子形成。
