Artificial microRNA-mediated resistance against the monopartite begomovirus Cotton leaf curl Burewala virus.
Artificial microRNA-mediated resistance against the monopartite begomovirus Cotton leaf curl Burewala virus.
复制标题
DOI：
10.1186/1743-422x-10-231
复制
发表时间：
2013-07-12
期刊：
影响因子：
4.8
通讯作者：
Mansoor S
中科院分区：
文献类型：
--
作者：
Ali I;Amin I;Briddon RW;Mansoor S
棉花卷叶病由 Begomovirus 属（双生病毒科）的单链 DNA 病毒引起，是巴基斯坦和印度西北部棉花种植的主要限制因素。目前只有具有中等耐受性的棉花品种才能对抗该病害。 microRNA (miRNA) 是一类内源性小 RNA 分子，在植物发育、信号转导以及对生物和生物胁迫的响应中发挥着重要作用。研究表明，可以对 miRNA 进行改造以改变其靶标特异性。这种人工 miRNA (amiRNA) 已被证明可以抵抗植物感染病毒。基于棉花 miRNA169a 序列，产生了两个含有 21 nt 棉花卷叶布雷瓦拉病毒 (CLCuBuV) V2 基因序列的 amiRNA 构建体，并转化到本塞姆塞姆氏烟草中。第一个 amiRNA 构建体 (P1C) 保留了 miR169a 序列，但替换的 21 nt 除外，而在第二个 (P1D) 中，改变了 miRNA169a 主链的序列以恢复成熟 miRNA 双链体的一些氢键。 P1C转基因植株在CLCuBV攻击下表现出良好的抗性；植物无症状，病毒 DNA 水平较低。对异源病毒的抵抗力较低，并且与amiRNA和V2基因序列之间的序列错配数量相关。 P1D 植物对所有测试病毒的攻击表现出整体较差的抵抗力。结果表明，amiRNA 方法可以在植物中提供针对单部分 Begomovirus 的有效抗性，并且具有广谱潜力，提供针对多种病毒的保护。此外，研究结果表明，耐药性水平取决于 amiRNA 与靶序列以及 miRNA 主链序列之间的互补水平，这与早期研究一致。
Cotton leaf curl disease, caused by single-stranded DNA viruses of the genus Begomovirus (family Geminiviridae), is a major constraint to cotton cultivation across Pakistan and north-western India. At this time only cotton varieties with moderate tolerance are available to counter the disease. microRNAs (miRNAs) are a class of endogenous small RNA molecules that play an important role in plant development, signal transduction, and response to biotic and a biotic stress. Studies have shown that miRNAs can be engineered to alter their target specificity. Such artificial miRNAs (amiRNAs) have been shown to provide resistance against plant-infecting viruses. Two amiRNA constructs, based on the sequence of cotton miRNA169a, were produced containing 21 nt of the V2 gene sequence of Cotton leaf curl Burewala virus (CLCuBuV) and transformed into Nicotiana benthamiana. The first amiRNA construct (P1C) maintained the miR169a sequence with the exception of the replaced 21 nt whereas in the second (P1D) the sequence of the miRNA169a backbone was altered to restore some of the hydrogen bonding of the mature miRNA duplex. P1C transgenic plants showed good resistance when challenge with CLCuBV; plants being asymptomatic with low viral DNA levels. The resistance to heterologous viruses was lower and correlated with the numbers of sequence mismatches between the amiRNA and the V2 gene sequence. P1D plants showed overall poorer resistance to challenge with all viruses tested. The results show that the amiRNA approach can deliver efficient resistance in plants against a monopartite begomoviruses and that this has the potential to be broad-spectrum, providing protection from a number of viruses. Additionally the findings indicate that the levels of resistance depend upon the levels of complementarity between the amiRNA and the target sequence and the sequence of the miRNA backbone, consistent with earlier studies.