Integrated morphological, phenolic, and transcript profiling reveals responses associated with colchicine-induced tetraploidy in Viola odorata L.

Abstract

Viola odorata L. (sweet violet) is an ornamental and medicinal species valued for its attractive flowers and diverse phenolic constituents. Although colchicine-induced polyploidization has been widely employed to improve ornamental and medicinal plants, the molecular responses associated with these improvements remain poorly characterized in V. odorata . Therefore, this study integrated morphological characterization, phenolic metabolite profiling, and gene expression analyses to investigate the effects of colchicine-induced tetraploidy. Seedlings at the four- to six-leaf stage were treated with colchicine (0–0.5%, w/v) by repeated application to the shoot apical meristem using cotton swabs for 24, 48, or 72 h. Stable tetraploid plants were confirmed by flow cytometry, with cytogenetic validation through chromosome counting following six months of greenhouse cultivation. Compared with diploid plants, tetraploids exhibited significantly enhanced vegetative and floral growth, including increased leaf area, petiole length, flower diameter, and biomass accumulation. Semi-quantitative RT-PCR indicated that the relative transcript abundance of PAL and Vok1 , estimated by densitometric analysis of PCR bands, was approximately 2.05- and 2.43-fold higher, respectively, in tetraploid than in diploid plants. HPLC analysis further revealed increased accumulation of several phenolic compounds, particularly p-coumaric acid (4.69-fold), together with higher levels of rutin, chlorogenic acid, rosmarinic acid, quercetin, cinnamic acid, caffeic acid, gallic acid, and apigenin. These findings indicate that integrating gene expression and phenolic metabolite profiling provides new insights into molecular responses associated with colchicine-induced tetraploidy in V. odorata , highlighting polyploidization as a promising strategy for modifying ornamental traits and phenolic metabolism.