Temporary inhibition of cell wall synthesis improves the transient expression of the GUS gene in Brassica napus mesophyll protoplasts

Transformation of Brassica napus mesophyll protoplasts was performed with the ß-glucuronidase gene fusion system. After electroporation, transient expression in protoplasts transformed directly after isolation was about 1 to 2 per million. By the use of 2,6-dichloro-benzonitrile, a non-toxic inhibitor of cell wall synthesis, and in the presence of 5% polyethyleneglycol, transformation of the cell material was performed three days after isolation. At that time, about 25–30% of the protoplasts had reached the first S-phase of the mitotic cycle. A 1000 fold increase of protoplasts expressing the ß-glucoronisidase gene transiently was obtained, in the partly synchronized protoplasts, compared to those transformed directly after isolation.Abbreviations CAT Chloroamphenicol acyltransferase - 2,6-DB 2,6-Dichlorobenzonitrile - EP-buffer electroporation buffer - GUS ßglucoronisidase - PEG Polyethylene glycol - PJ propidium iodide - Xgluc 5-bromo-4-chloro-3-indolyl glucoronide