Cassava geminivirus agroclones for virus-induced gene silencing in cassava leaves and roots

[en] Aim: We report the construction of a Virus-Induced Gene Silencing (VIGS) vector and an agroinoculation protocol for gene silencing in cassava (Manihot esculenta Crantz) leaves and roots. The African cassava mosaic virus isolate from Nigeria (ACMV-[NOg]), which was initially cloned in a binary vector for agroinoculation assays, was modified for application as VIGS vector. The functionality of the VIGS vector was validated in Nicotiana benthamiana and subsequently applied in wild-type and transgenic cassava plants expressing the uidA gene under the control of the CaMV 35S promoter in order to facilitate the visualization of gene silencing in root tissues. VIGS vectors were targeted to the Mg2+-chelatase gene in wild type plants and both the coding and promoter sequences of the 35S::uidA transgene in transgenic plants to induce silencing. We established an efficient agro-inoculation method with the hyper-virulent Agrobacterium tumefaciens strain AGL1, which allows high virus infection rates. The method can be used as a low-cost and rapid high-throughput evaluation of gene function in cassava leaves, fibrous roots and storage roots. Background: VIGS is a powerful tool to trigger transient sequence-specific gene silencing in planta. Gene silencing in different organs of cassava plants, including leaves, fibrous and storage roots, is useful for the analysis of gene function. Results: We developed an African cassava mosaic virus-based VIGS vector as well as a rapid and efficient agro-inoculation protocol to inoculate cassava plants. The VIGS vector was validated by targeting endogenous genes from Nicotiana benthamiana and cassava as well as the uidA marker gene in transgenic cassava for visualization of gene silencing in cassava leaves and roots. Conclusions: The African cassava mosaic virus-based VIGS vector allows efficient and cost-effective inoculation of cassava for high-throughput analysis of gene function in cassava leaves and roots. © 2018 The Author(s).

Disciplines :

Biotechnology

Author, co-author :

Lentz, E. M.; ETH Zurich-LFW, Department of Biology, Plant Biotechnology, E56.1, Universitaetstrasse 2, Zurich, 8092, Switzerland

Kuon, J.-E.; ETH Zurich-LFW, Department of Biology, Plant Biotechnology, E56.1, Universitaetstrasse 2, Zurich, 8092, Switzerland

Alder, A.; ETH Zurich-LFW, Department of Biology, Plant Biotechnology, E56.1, Universitaetstrasse 2, Zurich, 8092, Switzerland

Mangel, N.; ETH Zurich-LFW, Department of Biology, Plant Biotechnology, E56.1, Universitaetstrasse 2, Zurich, 8092, Switzerland

Zainuddin, I. M.; ETH Zurich-LFW, Department of Biology, Plant Biotechnology, E56.1, Universitaetstrasse 2, Zurich, 8092, Switzerland

McCallum, E. J.; ETH Zurich-LFW, Department of Biology, Plant Biotechnology, E56.1, Universitaetstrasse 2, Zurich, 8092, Switzerland

Anjanappa, R. B.; ETH Zurich-LFW, Department of Biology, Plant Biotechnology, E56.1, Universitaetstrasse 2, Zurich, 8092, Switzerland

Gruissem, W.; ETH Zurich-LFW, Department of Biology, Plant Biotechnology, E56.1, Universitaetstrasse 2, Zurich, 8092, Switzerland

Vanderschuren, Hervé ; Université de Liège - ULiège > Agronomie, Bio-ingénierie et Chimie (AgroBioChem) > Ingénierie des productions végétales et valorisation

Language :

English

Title :

Cassava geminivirus agroclones for virus-induced gene silencing in cassava leaves and roots

Publication date :

2018

Journal title :

Plant Methods

eISSN :

1746-4811

Publisher :

BioMed Central Ltd.

Volume :

Issue :

Peer reviewed :

Peer Reviewed verified by ORBi

Available on ORBi :

since 29 June 2019

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