Regeneration of Agrobacterium-mediated transgenic Brassica campestris ssp. parachinensis

Abstract

A procedure for the regeneration of transgenic Choy Sum (Brassica campestris ssp. parachinensis) is presented in this thesis. The protocol is based on infection of hypocotyl explants of 5-days old seedlings with Agrobacterium strain MOG301 carrying a disarmed binary vector pMOG410. The T-DNA region of this binary vector contains β-glucuronidase (gus) and neomycin phosphotransferase II (npt-II) as a reporter gene and a selectable marker gene. After co-cultivation for 48 hours, the hypocotyl explants were cultured on recovery medium containing 25 mg/L kanamycin for 1 to 2 weeks. The explants were subcultured in shooting medium containing 50 mg/L kanamycin, then subcultured every two weeks until shoot regenerated. Among nine cultivars tested, transformed shoots have been regenerated from 4 cultivars. These transformed shoots showed positive results in GUS histochemical assay. PCR and Southern blotting revealed that integration of the T-DNA region within the plant genome. The regeneration frequencies of transformants ranged from 0.83% to 5.26%. The transgenic Choy Sum all grew normally and developed fertile flowers.

Date of Award	1997
Original language	English
Awarding Institution	The Hong Kong University of Science and Technology