Multiple gene co-integration in Arabidopsis thaliana predominantly occurs in the same genetic locus after simultaneous in planta transformation with distinct Agrobacterium tumefaciens strains

A total of 243 transgenic T1-plant lines were produced by in planta transformation of Arabidopsis by a mixture of three distinct Agrobacterium tumefaciens strains carrying different plasmid constructs. PCR analysis indicated that about 30% of the transgenic plants were transformed with two different T-DNAs and additional 9.5% of the plants carried all genes from three different T-DNAs. Southern analysis revealed the integration from one to nine gene copies for each T-DNA used. Southern analysis of progenies in six lines displayed that the different transgenes were preferentially integrated into the same locus in a single plant genome. Analysis of fragment lengths, obtained in Southern analysis, allows to suppose the co-integration of several different T-DNAs in the single locus in direct, head-to-head or tail-to-tail orientation. PCR analyses of additional plants from these lines confirmed predominantly linked inheritance of all three transgenes integrated. About a half of T1-plants carrying all three transgenes were investigated by Northern hybridisations showing expressed mRNA at different levels. The expression of one transgene appears to have no effect on expression of the others and there is no correlation between the amount of mRNA and the number of gene copies in a single plant genome. The results reported here may be useful in plant metabolic engineering.