ArticlePlant methods2019
Highly efficient construction of infectious viroid-derived clones.
Article in Plant methods, 2019. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 4 papers.
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Who cites it
4 citing papers in PubMed, 19 citations in OpenAlex.
- A circular RNA vector for targeted plant gene silencing based on an asymptomatic viroid.The Plant journal : for cell and molecular biology · 2022Article
- Article
- Symptom Severity, Infection Progression and Plant Responses in Solanum Plants Caused by Three Pospiviroids Vary with the Inoculation Procedure.International journal of molecular sciences · 2021Article
- Hop stunt viroid: A polyphagous pathogenic RNA that has shed light on viroid-host interactions.Molecular plant pathology · 2021Article
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Authors and funding
4 authors at 1 institution in 1 country.
Funding
No grant is acknowledged in the PubMed record.
Abstract
backgroundViroid research generally relies on infectious cDNA clones that consist of dimers of the entire viroid sequence. At present, those dimers are generated by self-ligation of monomeric cDNA, a strategy that presents several disadvantages: (i) low efficiency, (ii) it is a non-oriented reaction requiring tedious screenings and (iii) additional steps are required for cloning into a binary vector for agroinfiltration or for in vitro RNA production.
resultsWe have developed a novel strategy for simultaneous construction of a viroid dimeric cDNA and cloning into a multipurpose binary vector ready for agroinfiltration or in vitro transcription. The assembly is based on IIs restriction enzymes and positive selection and supposes a universal procedure for obtaining infectious clones of a viroid independently of its sequence, with a high efficiency. Thus, infectious clones of one viroid of each family were obtained and its infectivity was analyzed by molecular hybridization.
conclusionThis is a zero-background strategy for direct cloning into a binary vector, optimized for the generation of infectious viroids. As a result, this methodology constitutes a powerful tool for viroid research and exemplifies the applicability of type IIs restriction enzymes and the lethal gene
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