Abstract: The frequency of shoot regeneration in autotetraploid broccoli was studied. Cotyledons with petiole as explants were cultured on MS media with different combinations of growth regulator. The effects of growth regulator combinations on induction of shoots were evaluated. The optimal media for inducting shoots were MS media containing 0.8% agar and 3% sucrose supplemented with 3.0 mg·L^-1 6-BA, 0.05 mg·L^-1 NAA. The shoot induction percentage was 63.3%. Then each shoot was transplanted on the MS medium containing 0.04 mg·L^-1 NAA and 4.0 mg·L^-1 6-BA proliferating about 6 times. The use of 0.1 mg·L^-1 NAA in the rooting medium increased the number of healthy roots per rooted plant (root ratio was 100%). In this study, we established high-efficient shoot regeneration system of autotetraploid broccoli and analyzed the ploidy stability of regenerates using flow cytometry. The result showed that the tetraploid relative DNA content was about two times of control. The identified result of flow cytometry according to the chromosome analysis implied the regeneration plantlets keep the original ploidy.

Key words: broccoli, autotetraploid, cotyledons with petiole, in vitro culture, flow cytometry machine