Establishment of a versatile virus-based system for elucidating herbicide resistance in Galium aparine

Herbicide resistance challenges sustainable weed management in crop fields. However, the lack of gene manipulation tools for weeds hinders studies of herbicide resistance evolution and regulation. Here, we identified that tobacco mosaic virus (TMV) can systemically infect Galium aparine, a noxious broadleaf weed common in wheat and rapeseed fields, among 13 broadleaf weeds. The TMV-based expression system can be used to elucidate herbicide resistance in G. aparine. We demonstrated that virus-based expression of BAR, ALS, and its site-directed mutagenized form in G. aparine plants can impart herbicide resistance. To expand the application of this system in metabolic resistance studies, 10 upregulated P450 genes in the tribenuron-methyl-resistant G. aparine population plants were identified. Expression of these P450 genes in G. aparine via this system enhanced tribenuron-methyl resistance in G. aparine plants. Additionally, we confirmed Eleusine indica CYP81A104 functions in herbicide resistance through heterologous expression in G. aparine plants, suggesting that this system can be used to investigate the roles of genes from other weed species. The TMV vector-based assay system enabled the functional validation of weed genes within 2 months, which is shorter than the time required in other heterologous expression systems (ie, stable expression of weed genes in rice or Arabidopsis). Thus, this virus-based system provides insights into target-site and/or metabolic-based resistance mechanisms to herbicides in G. aparine and possibly in other weed species.