J Integr Plant Biol. ›› 1999, Vol. 41 ›› Issue (7): -.

• Research Articles •    

Genetic Transformation of Protoplasts from Peucedanum terebinthaceum and Regeneration of Herbicide resistant Plantlets

WANG Xiu-Feng, YANG Ai-Fang and ZHANG Ju-Ren   

Abstract: Calli produced from the segments of young petioles of Peucedanum terebinthaceum Fisch. ex Turcz. were transfered into liquid medium for suspension culture. The protoplasts were isolated from cells of clusters by enzymes digestion. The mutant als ( acetolactate synthase) gene of Arabidopsis thaliana ( L. ) Heynh. Was transferred into protoplasts of P. terebinthaceum with PEG method. A transformation medium containing 40 - 50 μg/mL DNA and 10% PEG and an incubation condition of 26 ℃ for 25 minutes in dark revealed the best result. Supplementation with 0~60μg/mL ctDNA into the medium gave no remarkable effect on transformation frequency. After cell clumps formed from the treated protoplasts in liquid medium, they were transferred onto proliferation medium containing 0.01 mg/L chlorsulfuron. The resistant clones grew 20 days later, and were transferred onto differentiation medium containing 0.03 mg/L chlorsulfuron. Plantlets regenerated via embryoid pathway. Southern blot hybridization with the 32p labeled 2.5 kb fragments of p35s- als demonstrated that the mutant als gene had inserted into P. terebinthaceum genome. Chlorsulfuron-resistant calli which grew normally were produced when segments of tmnsgenic plantlets were cultured on the proliferation medium containing 0.01 mg/L chlorsulfuron, whereas the growth of untransgenic explants were inhibited. Meanwhile, the transgenic plants could stand high density of chlorsulfuron up to 0.06 mg/L, but not the wild plants. These results indicated that the exogenous als gene had integrated into P. terebinthaceum genome and could transcript and translate stably.

Key words: Protoplast, Genetic transformation, als gene, chlorsulfuron, Peucedanum terebinthaceum

Editorial Office, Journal of Integrative Plant Biology, Institute of Botany, CAS
No. 20 Nanxincun, Xiangshan, Beijing 100093, China
Tel: +86 10 6283 6133 Fax: +86 10 8259 2636 E-mail: jipb@ibcas.ac.cn
Copyright © 2022 by the Institute of Botany, the Chinese Academy of Sciences
Online ISSN: 1744-7909 Print ISSN: 1672-9072 CN: 11-5067/Q