Effect of Temperature on Microspore Embryogenesis and Regeneration of Doubled Haploid Plants in Three Canola (Brassica napus L.) Hybrids

M. Oroojloo1,; M. E. Shariatpanahi,; S. Habibzadeh,; M. Emamifar,; F. Javidfar,

doi:10.22092/spij.2017.111057

Effect of Temperature on Microspore Embryogenesis and Regeneration of Doubled Haploid Plants in Three Canola (Brassica napus L.) Hybrids

Authors

10.22092/spij.2017.111057

Abstract

In the present study microspore embryogenesis and regeneration were investigated in canola (Brassica nupus L.) to develop doubled haploid plants. Canola cultivars Hyola 420, Hyola 401 and F1 hybrid RGS003 ´ ARC5 were used as plant materials. Microspores at late unicellular- early bicellular developmental stages were isolated from flower buds with an approximate length of 2.5-3.5 mm and cultured in NLN medium containing 13% sucrose. Isolated microspores were stressed with heat shock in different durations and temperatures including 37°C for 8 hours, 35 °C for 18 hours, 32°C for 72 hours and 30 °C for 10 and 14 days. Microspore-derived embryos were induced only when heat shock treatment of 30oC for 10-14 days was applied, although 30oC for 14 days showed better embryogenesis and regeneration. For blocking the secondary embryogenesis, embryos in cotyledonery stage were transferred to B5 medium supplemented by GA3 (0.01 mgl-1) in dark condition for 10 day at 4 oC and then transferred to the light condition at 25 oC to B5 medium without GA3. For doubling the ploidy level of haploids and production of doubled haploids, plantlets were treated with colchicine (0.34%) for 1.5 hours. Doubled haploid seeds were collected from all three genotypes tested.

Keywords