In planta transformation of Dendrobium nobile by ovary-injection of Agrobacterium.

Dendrobium nobile was transformed by ovary-injection of Agrobacterium.After artificial pollination in 10,20,30,40,45,50 and 60 d,EHA105(pCAMBIA1301) harboring both β-glucuronidase gene(GUS) and hpt was injected into the ovary.The seeds of split fruit of D.nobile were cultured on the N16 culture medium by aseptic processing and detected by GUS expression.Forty-five days after cultured,the protocorms were diverted to N16 medium with hygromycin to gain hygromycin-resistance protocorms,then four randomly selected hygromycin-resistance protocorms were detected by PCR analysis.The study showed that ovary-injection of agrobacterium had harmful effects on the fruit development and seed formation.Ovary-injection of agrobacterium after artificial pollination 45 d had the highest efficiency,18% seeds from the injected position showing GUS expression.Eleven hygromycin-resistant plants were gained in this study,and four randomly selected hygromycin-resistant plants were detected by PCR analysis and all of them were positive.The results showed that ovary-injection of agrobacterium transformation had advantages in genetic transformation of orchids.