Hairpin RNA Targeting Multiple Viral Genes Confers Strong Resistance to Rice Black-Streaked Dwarf Virus.
Hairpin RNA Targeting Multiple Viral Genes Confers Strong Resistance to Rice Black-Streaked Dwarf Virus.
复制标题
靶向多个病毒基因的发夹RNA赋予水稻黑条矮缩病毒强大的抗性
DOI：
10.3390/ijms17050705
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发表时间：
2016-05-11
影响因子：
5.6
通讯作者：
Yang J
中科院分区：
文献类型：
--
作者：
Wang F;Li W;Zhu J;Fan F;Wang J;Zhong W;Wang MB;Liu Q;Zhu QH;Zhou T;Lan Y;Zhou Y;Yang J
水稻黑条矮缩病毒（RBSDV）属于呼肠孤病毒科斐济病毒属，在亚洲水稻产区造成严重的产量损失。RNA沉默作为植物抗病毒的一种天然防御机制，已被成功地用于植物（包括水稻）的抗病毒工程。在这项研究中，我们产生了转基因水稻株系窝藏发夹RNA（hpRNA）构建针对四个RBSDV基因，S1，S2，S6和S10，编码RNA依赖的RNA聚合酶，推定的核心蛋白，RNA沉默抑制子和外壳蛋白，分别。田间育苗和人工接种试验均表明，转基因株系对RBSDV具有较强的抗性。分离的转基因群体中的RBSDV抗性与hpRNA转基因的存在完全相关。此外，hpRNA转基因在高抗性转基因株系中表达，产生丰富水平的21-24 nt小干扰RNA（siRNA）。通过小RNA深度测序，RBSDV抗性转基因系检测到hpRNA转基因中所有四个病毒基因序列的siRNA，这表明整个嵌合融合序列可以被Dicer有效加工成siRNA。综上所述，我们的研究结果表明，靶向多个病毒基因的长hpRNA可用于在水稻以及其他植物物种中产生稳定和持久的病毒抗性。
Rice black-streaked dwarf virus (RBSDV) belongs to the genus Fijivirus in the family of Reoviridae and causes severe yield loss in rice-producing areas in Asia. RNA silencing, as a natural defence mechanism against plant viruses, has been successfully exploited for engineering virus resistance in plants, including rice. In this study, we generated transgenic rice lines harbouring a hairpin RNA (hpRNA) construct targeting four RBSDV genes, S1, S2, S6 and S10, encoding the RNA-dependent RNA polymerase, the putative core protein, the RNA silencing suppressor and the outer capsid protein, respectively. Both field nursery and artificial inoculation assays of three generations of the transgenic lines showed that they had strong resistance to RBSDV infection. The RBSDV resistance in the segregating transgenic populations correlated perfectly with the presence of the hpRNA transgene. Furthermore, the hpRNA transgene was expressed in the highly resistant transgenic lines, giving rise to abundant levels of 21–24 nt small interfering RNA (siRNA). By small RNA deep sequencing, the RBSDV-resistant transgenic lines detected siRNAs from all four viral gene sequences in the hpRNA transgene, indicating that the whole chimeric fusion sequence can be efficiently processed by Dicer into siRNAs. Taken together, our results suggest that long hpRNA targeting multiple viral genes can be used to generate stable and durable virus resistance in rice, as well as other plant species.