Division of Nematology, ICAR-Indian Agricultural Research Institute, New Delhi, India.
ICAR-National Institute for Plant Biotechnology, New Delhi, India.
Pest Manag Sci. 2021 Jul;77(7):3396-3405. doi: 10.1002/ps.6384. Epub 2021 Apr 13.
Occurrence of multiple biotic stresses on crop plants result in drastic yield losses which may have severe impact on the food security. It is a challenge to design strategies for simultaneous management of these multiple stresses. Hence, establishment of innovative approaches that aid in their management is critical. Here, we have introgressed a micro RNA-induced gene silencing (MIGS) based combinatorial gene construct containing seven target gene sequences of cotton leaf curl disease (CLCuD), cotton leaf hopper (Amrasca biguttula biguttula), cotton whitefly (Bemisia tabaci) and root-knot nematode (Meloidogyne incognita).
Stable transgenic lines of Nicotiana benthamiana were generated with the T-DNA harboring Arabidopsis miR173 target site fused to fragments of Sec23 and ecdysone receptor (EcR) genes of cotton leaf hopper and cotton whitefly. It also contained C2/replication associated protein (C2/Rep) and C4 (movement protein) along with βC1 gene of betasatellite to target CLCuD, and two FMRFamide-like peptide (FLP) genes, Mi-flp14 and Mi-flp18 of M. incognita. These transgenic plants were assessed for the amenability of MIGS approach for pest control by efficacy evaluation against M. incognita. Results showed successful production of small interfering RNA (siRNA) through the tasiRNA (trans-acting siRNA) pathway in the transgenic plants corresponding to Mi-flp18 gene. Furthermore, we observed reduced Mi-flp14 and Mi-flp18 transcripts (up to 2.37 ± 0.12-fold) in females extracted from transgenic plants. The average number of galls, total endoparasites, egg masses and number of eggs per egg mass reduced were in the range 27-62%, 39-70%, 38-65% and 34-49%, respectively. More importantly, MIGS transgenic plants showed 80% reduction in the nematode multiplication factor (MF).
This study demonstrates successful validation of the MIGS approach in the model plant, N. benthamiana for efficacy against M. incognita, as a prelude to translation to cotton. © 2021 Society of Chemical Industry.
作物受到多种生物胁迫的影响会导致严重减产，这可能对粮食安全产生严重影响。设计同时管理这些多种胁迫的策略是一项挑战。因此，建立有助于管理它们的创新方法至关重要。在这里，我们导入了一个基于 micro RNA 诱导的基因沉默（MIGS）的组合基因构建体，该构建体包含棉花曲叶病（CLCuD）、棉叶蝉（Amrasca biguttula biguttula）、棉粉虱（Bemisia tabaci）和根结线虫（Meloidogyne incognita）的七个靶基因序列。
用含有棉叶蝉和棉粉虱 Sec23 和蜕皮激素受体（EcR）基因的 Arabidopsis miR173 靶位点融合片段的 T-DNA 生成了稳定的 Nicotiana benthamiana 转基因系。它还包含 C2/复制相关蛋白（C2/Rep）和 C4（运动蛋白）以及 betasatellite 的βC1 基因，以靶向 CLCuD，以及两个 FMRFamide 样肽（FLP）基因 Mi-flp14 和 Mi-flp18 来自 M. incognita。通过对 M. incognita 的功效评估，评估了这些转基因植物对 MIGS 方法进行害虫防治的适宜性。结果表明，在对应于 Mi-flp18 基因的转基因植物中，通过 tasiRNA（反式作用 siRNA）途径成功产生了小干扰 RNA（siRNA）。此外，我们观察到从转基因植物中提取的雌性中 Mi-flp14 和 Mi-flp18 转录本减少（高达 2.37±0.12 倍）。根结数、总内寄生蜂、卵块和每卵块的卵数分别减少了 27-62%、39-70%、38-65%和 34-49%。更重要的是，MIGS 转基因植物中的线虫繁殖因子（MF）减少了 80%。
这项研究证明了 MIGS 方法在模式植物 Nicotiana benthamiana 中对 M. incognita 的有效性，为向棉花的转化奠定了基础。 © 2021 化学工业协会。
