Transgenic flax plants from Agrobacterium mediated transformation: incidence of chimeric regenerants and inheritance of transgenic plants

Abstract

Transgenic flax ( Linum usitatissimum) plants were generated by Agrobacterium tumefaciens mediated gene transfer. The binary plasmid p35SGUSINT in the octopine type A. tumefaciens strain GV2260 was used as the marker/vector system. An intron-containing β-glucoronidase (GUS) gene driven by the cauliflower mosaic virus 35S promoter served as the reporter, detected by the histochemical assay. The neomycin phosphotransferase II gene in the T-DNA provided resistance to kanamycin and was used as the selectable marker. Using the histochemical GUS assay, differentially stained tissues were observed in the stems and leaves of the selected shoots at a high frequency. When the selected plants were allowed to self-cross and set seeds, approximately 45% of the lines showed much lower transgenic segregation ratios than expected of Mendelian inheritance, or did not produce any transgenic offspring, even though F 0 plants passed both the histochemical assay and polymerase chain reaction test for the presence and expression of transgenes. We suggest that those transgenic plants with irregular transgene expression are often chimeras consisting of transformed and non-transformed tissues, which are the result of the multicellular origin of shoot organogenesis from flax hypocotyls, with efficient protection from the selection agent of non-transformed cells by transformed ones. Non-chimeric transgenic plants were successfully recovered from transformed cells in chimeric plants using internode propagation in vitro and/or analysis of a large progeny population. This transformation system could be used both by those desiring chimeric plants (e.g. for developmental studies) or those desiring to recover non-chimeric transgenic plants.