INDUCTION OF MICROSPORE-DERIVED EMBRYOS OF BRASSICA-NAPUS L WITH POLYETHYLENE-GLYCOL (PEG) AS OSMOTICUM IN A LOW SUCROSE MEDIUM

Isolated microspores of Brassica napus were cultured on high concentra tions of mannitol or polyethylene glycol (PEG 4000), with only a Very limited amount of sucrose (0.08-0.1%) provided as carbohydrate source in the medium. While microspores cultured on high mannitol yielded no embryos and no embryogenic cell divisions were observed, microspores o n high PEG developed into embryos within 2 weeks, and the embryo yield appeared comparable to that of the sucrose control. When placed under light, PEG embryos quickly changed color from yellow to dark green, w hile sucrose embryos first remained yellowish and then slowly changed color to pale green. Three-week-old PEG embryos were strikingly simila r to immature zygotic embryos developed in ovulo, dissected at 14-15 d ays post-anthesis (DPA), while sucrose embryos differed from the latte r in the size and shape, color and morphology of their cotyledons. The se results demonstrate that in microspore embryogenesis of Brassica na pus: (1) the level of metabolizable carbohydrate required for microspo re embryo induction and formation appears to be substantially less tha n commonly used amounts, (2) sucrose as an osmoticum can be replaced w ith high-molecular-weight PEG. With further improvement the new method described here might be suitable for other Brassica species and would have a great potential application in breeding programs.