Integrated phenotypic and transcriptomic analysis of herbicide stress response in hemp

Abstract Hemp ( Cannabis sativa L.) faces significant weed management challenges due to limited registered herbicides. This study evaluated tolerance of hemp to eight post-emergence herbicides and investigated the molecular mechanisms underlying tolerance to bispyribac-sodium through RNA sequencing. Greenhouse screening revealed that ACCase inhibitors caused minimal phytotoxicity, while Atrazine, Metribuzin, metsulfuron-methyl, and oxyfluorfen caused complete plant mortality. Bispyribac-sodium demonstrated partial tolerance with a GR 50 of 21.55 g a.i. ha − 1 , slightly above the field-recommended dose. Transcriptomic profiling revealed significant differential gene expression at 24 and 48 h after treatment, with early responses characterized by activation of MAPK signaling, secondary metabolite biosynthesis, and xenobiotic detoxification, while later responses shifted toward protein quality control and endoplasmic reticulum stress. A core set of 21 consistently upregulated detoxification genes, including tau-class glutathione S-transferases, UDP-glycosyltransferases, cytochrome P450s (CYP71, CYP72, CYP81, and CYP716 families), and an ABCB transporter, implicate a coordinated phase I-III detoxification framework in hemp’s herbicide tolerance. Weighted gene co-expression network analysis identified the yellow, purple, and pink modules as candidate co-expression networks underlying this response. These findings provide the first molecular reference for herbicide response in hemp and an evidence-based foundation for weed management strategies.