English

Abstract

Two germinating seed transformation methods i.e., scarifying and puncturing methods, were used to transfer the bar gene into an inbred maize line. We generated transgenic glufosinate-tolerant maize plants, and conducted a comparison of the two transformation methods. We used the plasmid pCAMBAR.CHI.11 as the donor and germinating maize seeds as the recipients. The germinating seeds were subjected to puncture or scarifying treatment after being soaked in water for 12 h. The injured seeds were co-cultivated with Agrobacterium for 24 h and then sown in the experimental plots. Putative transgenic maize plants were selected by basta tolerance screening and PCR detection, and were further confirmed by Southern blotting. Following glufosinate tolerance assay and molecular detection of transgenic lines of the transgenic seedling (T0), first generation of transgenic seedling (T1) and second generation of transgenic seedling (T2), we have confirmed that the bar gene was stably inherited and expressed. The PCR amplification result for T2 transgenic plants showed that the genetic segregation ratio of the bar gene followed the 3:1 ratio of a single dominant Mendelian factor. The study proved that both the germinating seed treatment approaches are rapid and simple plant transformation methods. In particular, the application of the puncture method can expand the tissue culture free transformation to dicotyledonous plants. © 2016 Friends Science Publishers