Department of Biosciences, Jamia Millia Islamia (Central University), Jamia Nagar, New Delhi, 110025, India.
Department of Molecular Medicine, Jamia Hamdard, Hamdard Nagar, New Delhi, 110062, India.
Sci Rep. 2021 Jul 8;11(1):14172. doi: 10.1038/s41598-021-93502-3.
Cotton leaf curl disease (CLCuD), caused by begomoviruses in combination with betasatellite molecule, has adversely affected cotton industry of Indian subcontinent. To devise a CLCuD-control strategy, RNAi-mediated approach was followed in this study. Gossypium hirsutum cv. HS6 plants were transformed with intron-hairpin RNAi (ihpRNAi-C4) construct carrying silencing suppressor C4 gene of Cotton leaf curl Multan virus (CLCuMuV). Efficacy of the construct in imparting CLCuD resistance was evaluated in transgenic (T, T) cotton lines. Accumulation of CLCuMuV/betasatellite and attenuation of CLCuD symptoms in the transgenic lines were monitored at different times interval after virus inoculation. Northern hybridization revealed the expression of C4-gene derived siRNA. Expression of the ihpRNAi transcript was recorded higher in transgenic lines expressing siRNA which supposedly targeted the C4 gene. A significant delay in detection of virus as well as betasatellite was observed in the transgenic lines. At 30 days post inoculation (dpi), none of the lines tested positive. At 45 dpi, however, it could be detected in few lines having much lower titre as compared to non-transformed control plants. Notably, till 60 dpi, no significant progression of the virus/betasatellite DNA was observed and the plants did not exhibit any characteristic CLCuD symptoms. A tolerance phenomenon leading to escape of CLCuD symptoms in the transformed cotton was described.
棉花曲叶病（CLCuD）是由伴随β卫星分子的双生病毒引起的，对印度次大陆的棉花产业造成了不利影响。为了制定 CLCuD 防控策略，本研究采用了 RNAi 介导的方法。用携带沉默抑制子 C4 基因的棉花曲叶病 Multan 病毒（CLCuMuV）的内含子发夹 RNAi（ihpRNAi-C4）构建体转化了陆地棉品种 HS6 植株。在转基因（T、T）棉花株系中评估了该构建体赋予 CLCuD 抗性的效果。在病毒接种后不同时间间隔监测了 CLCuMuV/β卫星的积累和 CLCuD 症状的减轻。Northern 杂交显示了 C4 基因衍生的 siRNA 的表达。在表达 siRNA 的转基因株系中记录到 ihpRNAi 转录物的表达更高，推测该 siRNA 靶向 C4 基因。在转基因株系中观察到病毒和β卫星的检测明显延迟。在接种后 30 天（dpi），没有检测到任何阳性株系。然而，在 45 dpi 时，与未转化对照植物相比，在少数株系中检测到的病毒载量要低得多。值得注意的是，直到 60 dpi，都没有观察到病毒/β卫星 DNA 的显著进展，并且这些植物没有表现出任何典型的 CLCuD 症状。描述了一种导致转化棉中 CLCuD 症状逃避的耐受性现象。
