Agrobacterium-mediated transformation of Asparagus officinalis L.: molecular and genetic analysis of transgenic plants

Four long-term embryogenic lines of Asparagus officinalis were co-cultured with the hypervirulent Agrobacterium tumefaciens strain AGL1Gin carrying a uidA gene and an nptII gene. 233 embryogenic lines showing kanamycin resist ance and beta -glucuronidase (GUS) activity were obtained. Transformation f requencies ranged from 0.8 to 12.8 transformants per gram of inoculated som atic embryos, depending on the line. Southern analysis showed that usually 1 to 4 T-DNA copies were integrated. Regenerated plants generally exhibited the same insertion pattern as the corresponding transformed embryogenic li ne. T-1 progeny were obtained from crosses between 6 transformed plants con taining 3 or 4 T-DNA copies and untransformed plants. They were analysed fo r GUS activity and kanamycin resistance. In three progenies, Mendelian 1:1 segregations were observed, corresponding to one functional locus in the pa rent transgenic plants. Southern analysis confirmed that T-DNA copies were inserted at the same locus. Non-Mendelian segregations were observed in the other three progenies. T-2 progeny also exhibited non-Mendelian segregatio ns. Southern analysis showed that GUS-negative and kanamycin-sensitive plan ts did not contain any T-DNA, and therefore inactivation of transgene expre ssion could not be responsible for the abnormal segregations.