<i>Agrobacterium tumefaciens</i>‐mediated barley transformation

TLDR

Barley was transformed by infecting immature embryo explants with Agrobacterium tumefaciens carrying a binary vector containing bar and gus, followed by selection of bialaphos‑resistant cultures and plant regeneration. From 1282 embryos, 54 independent lines were obtained (4.2 % efficiency); transgene copy numbers ranged from single to ≥10, 16 of 18 mature plants were fertile, and both bar and gus were expressed and co‑segregated, generally following Mendelian patterns except in one family where two unlinked loci were identified.

Abstract

Genetically transformed barley was produced by eco‐cultivating immature embryo explants with Agrobacterium tumefaciens carrying a binary vector coding for chimaeric bacterial genes, bar and gus , and selecting for bialaphos‐resistant cultures from which plants were regenerated. Integration of both genes was confirmed by gel blot hybridization analysis of DNA from the transformed plants and their progenies. From 1282 embryos, plants were recovered for 54 independently transformed lines, giving a transformation efficiency of 4.2%. Transgene numbers in the different lines ranged from single copy insertion to at least ten copies. Sixteen out of 18 plants grown to maturity were fully fertile. Both marker genes, bar and gus , were expressed and co‐segregated in the T 1 progeny plants. In the majority of cases, the genes showed Mendelian segregation predicted for transgene insertion at a single locus. In one family with multiple transgene insertions, molecular analysis of T 1 and T 2 plants suggested that the T‐DNA had inserted at two unlinked loci.