What is a repositorium?

The repositorium is a searchable database that provides data on relevant articles from journals, company web pages and web pages of governmental agencies about studies/applications of genome-editing in model plants and agricultural crops in the period January 1996 to May 2018. Search options are article type, technique, plant, traits or free text. The repositorium is based on the systematic map of Dominik Modrzejewski et al., published in the journal environmental evidence. (Download article PDF).

Multiplexed Gene Editing and Protein Overexpression Using a Tobacco mosaic virus Viral Vector

Typ / Jahr

Journal Article / 2017


Cody, Will B.; Scholthof, Herman B.; Mirkov, T. Erik


Development of CRISPR/Cas9 transient gene editing screening tools in plant biology has been hindered by difficulty of delivering high quantities of biologically active single guide RNAs (sgRNAs). Furthermore, it has been largely accepted that in vivo generated sgRNAs need to be devoid of extraneous nucleotides, which has limited sgRNA expression by delivery vectors. Here, we increased cellular concentrations of sgRNA by transiently delivering sgRNAs using a Tobacco mosaic virus-derived vector (TRBO) designed with 5' and 3' sgRNA proximal nucleotide-processing capabilities. To demonstrate proof-of-principle, we used the TRBO-sgRNA delivery platform to target GFP in Nicotiana benthamiana (16c) plants, and gene editing was accompanied by loss of GFP expression. Surprisingly, indel (insertions and deletions) percentages averaged nearly 70% within 7 d postinoculation using the TRBO-sgRNA constructs, which retained 5' nucleotide overhangs. In contrast, and in accordance with current models, in vitro Cas9 cleavage assays only edited DNA when 5' sgRNA nucleotide overhangs were removed, suggesting a novel processing mechanism is occurring in planta. Since the Cas9/TRBO-sgRNA platform demonstrated sgRNA flexibility, we targeted the N. benthamiana NbAGO1 paralogs with one sgRNA and also multiplexed two sgRNAs using a single TRBO construct, resulting in indels in three genes. TRBO-mediated expression of an RNA transcript consisting of an sgRNA adjoining a GFP protein coding region produced indels and viral-based GFP overexpression. In conclusion, multiplexed delivery of sgRNAs using the TRBO system offers flexibility for gene expression and editing and uncovered novel aspects of CRISPR/Cas9 biology.

CRISPR-Cas Systems; Gene Editing/methods; tobacco; Tobacco Mosaic Virus
Plant physiology
Periodical Number
Page range


ID Corresponding Author
Plant Species GE Technique
Sequence Identifier
Type of Alteration
Progress in Research
Key Topic
1190 Scholthof, Herman B.; Mirkov, T. Erik
Nicotiana benthamiana CRISPR/Cas9
Green fluorescence
Basic research
Basic research
1191 Scholthof, Herman B.; Mirkov, T. Erik
Nicotiana benthamiana CRISPR/Cas9
Role in RNA silencing
Basic research
Basic research