A review of factors affecting the success of geminivirus infectious clones.

Key message: In this review, we provide a summary of factors that affect the successful infection of geminivirus clones in plants to enable the greater understanding of plant-virus interactions. Geminiviruses are single-stranded DNA viruses that can cause significant losses in economically important crops worldwide. Considerable efforts have been made to study the geminiviruses in detail, which has resulted in the construction of many infectious clones for the vast diversity of geminiviruses. In laboratory conditions, agrobacterium or occasionally biolistic methods are used to deliver viral DNA to the plant cell. However, not every delivered viral DNA will develop into an infection due to several reasons. In this manuscript, we review the factors that affect the success of geminivirus infectious clones. Factors affecting virus infection including the methods of inoculating in vitro-generated viral DNA constructs are often neglected, leading to failed virus infections and drawing wrong conclusions. Deciding exactly where on the plant to inoculate, what age of plant, and what agrobacterium strain are all examples of variables which may influence an infection. We find that stem injections of agrobacterium into young seedlings with an optical density at 600 nm (OD₆₀₀) in the 0.1-0.3 range are an optimal starting point for studies. This review will provide a thorough compilation of inoculation methods and use this to discuss the deeper mechanisms at play during the initial infection of plants with geminivirus infectious clones.