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

Plant Methods
Ezequiel Matias LentzHervé Vanderschuren

Abstract

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. VIGS is a powerful tool to trigger transient sequence-specific gene silencing in planta. Gene silencing i...Continue Reading

References

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Citations

Jun 11, 2020·Annual Review of Virology·Peter AbrahamianJohn Hammond
Aug 25, 2019·International Journal of Molecular Sciences·Hongqiu ZengHaitao Shi
Jun 6, 2019·Methods and Protocols·Ezequiel Matías LentzHervé Vanderschuren
Jun 20, 2020·The Journal of Biological Chemistry·Teresa B Fitzpatrick, Lottie M Chapman
Jan 20, 2022·Horticulture Research·Lucky PaudelChristopher I Cazzonelli

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Datasets Mentioned

BETA
AF234297.1

Methods Mentioned

BETA
transgenic
nucleic acid extraction

Software Mentioned

CIAT
Adobe Photoshop

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