A simple widely applicable hairy root transformation method for gene function studies in medicinal plants.

Genetic transformation is a fundamental tool in molecular biology research of medicinal plants. Tailoring transgenic technologies to each distinct medicinal plant would necessitate a substantial investment of time and effort. Here, we present a simple hairy root transformation method that does not require sterile conditions, utilizing strain K599 and the visible RUBY reporter system. Transgenic hairy roots were obtained for six tested medicinal plant species, roots or rhizomes of which have recognized medicinal value, spanning four botanical families and six genera (, , , , , and ). Furthermore, two previously identified UGTs that convert liquiritigenin into liquiritin in heterologous systems were studied using the method. Our results indicate that overexpression of but not and Cas9-mediated knockout of profoundly influenced the accumulation of liquiritin and isoliquiritin in licorice roots. Therefore, the method described here represents a simple, rapid and widely applicable hairy root transformation method that enables fast gene functional study in medicinal plants.