Development of an efficient selection procedure for high imperatorin-yielding hairy root clones of Urena lobata L. using morphological and molecular markers

• Published in: Plant cell, tissue and organ culture tissue and organ culture, 2024-11, Vol.159 (2), p.35, Article 35
• Main Authors: Cao, Dai Minh, Bui, Anh Lan, Van Bui, Le, Quach, Phuong Ngo Diem
• Format: Article
• Language: English
• Subjects: Agrobacterium rhizogenes; Biomedical and Life Sciences; Chemical synthesis; Hairy root; Life Sciences; Liquid culture; Medicinal plants; Morphology; Original Article; Plant Genetics and Genomics; Plant layout; Plant Pathology; Plant Physiology; Plant Sciences; Roots; Urena lobata
• ISSN: 0167-6857; 1573-5044
• DOI: 10.1007/s11240-024-02898-z

The root of Urena lobata L. has been shown to be able to synthesize imperatorin (a furocoumarin), which is a medicinal compound with anti-inflammatory and anticancer effects. Here, we describe for the first time the production of the hairy roots (HRs) of this plant via explant cocultivation with the ATCC 15,834 strain of Agrobacterium rhizogenes to produce imperatorin. After screening 138 h clones, C4 and E3 were determined to be high-yield root clones because their dry growth index (DGI) and imperatorin content were markedly higher than others with 33.6 and 0.078 µg g − 1 DW, respectively. To develop an efficient selection procedure for high imperatorin-yielding HR clones, several morphological and molecular parameters of C4 and E3 were evaluated. These clones were characterized by their root length, number of lateral roots/main root, diameter of the main root, and diameter of the lateral root greater than 20 roots, 11 cm, 0.07 mm, and 0.04 mm, respectively. Because of the eventuality of transformation, we hypothesized that these high-yield clones may include specific molecular sequences associated with growth and imperatorin production. Molecular markers of C4 and E3 were simultaneously confirmed by amplifying a 600-bp fragment of the AGAC4 primer using ISSR technique. A selection procedure was then developed included four steps: (I) Induction to create various HR clones; (II) Selection of clones with morphological markers after 21 days of induction; (III) Selection of clones with molecular markers after 21 days of induction; (IV) Selection of high-yield clones with DGI and imperatorin content after 28 days in liquid culture. The developed procedure could increase the selection efficiency to 89.2% while also reducing the number of screened root clones to 4.7-fold. Furthermore, the selected high-yield HR clones grew and accumulated imperatorin in a sustainable manner for the subsequent three subcultures. Key message The HRs of Urena lobata were generated for the production of imperatorin and the efficiency for the selection of high imperatorin-yielding root clones was increased by using morphological and molecular markers.