Preliminarily chloroplast transformation of tobacco variety V2 by bombardment

Abstract

V2 is a high yield, large tobacco variety which is cultivated in many material areas of Vietnam. For successful chloroplast transformation, an efficient shoot regeneration system was required. Highest frequent regeneration was obtained when culturing V2 leaf discs on the basis medium supplemented NAA 0.1 mg/l and BA 1mg/l. Besides, the key factor in determining transgenic plants is consistently selective pressure. V2 variety of tobacco was sensitive to the antibiotics spectinomycin and streptomycin. At the concentration of 80 mg/l, spectinomycin inhibited completely shoot regeneration and plant growth.  With streptomycin, the threshold is 125 mg/l. HIV-1 p24 and aadA genes were transferred to chloroplasts of the tobacco V2 variety by bombardment. Putatively transgenic plants were capable of resisting to streptomycin and spectinomycin up to 500 mg/l concentration. Through initial PCR testing, HIV-1 p24 was assumed present in the genome of the transgenics with the specific amplification band of 700 bp.