Cotyledons:a useful biological material for transient gene expression analysis in rapeseed(Brassica napus L.)

Polyethylene glycol (PEG)-mediated DNA transformation for transient gene expression in protoplasts and Agrobacterium tumefaciens-mediated transformation in lower epidermis of leaves are readily available in several plant species. In the study, these two versatile tools were used in rapeseed. A simple and efficient method was established for isolating protoplasts from rapeseed cotyledons and leaves, and found that cotyledons might be better than true leaves. Transient expression analysis showed that yellow fluorescent protein (YFP) and luciferase (LUC) could be expressed in rapeseed protoplasts. Moreover,GUS histochemical assays indicated that Agrobacterium-mediated DNA transient expression was achievable only in lower epidermis of rapeseed cotyledons and expression signal was the highest on the 5th day after injection with the bacterial suspension (OD600=0.8).These methods might provide valuable tools for rapid functional gene analysis in rapeseed.