The T-DNA integration pattern in Arabidopsis transformants is highly determined by the transformed target cell.
Transgenic loci obtained after Agrobacterium tumefaciens-mediated transformation can be simple, but fairly often they contain multiple T-DNA copies integrated into the plant genome. To understand the origin of complex T-DNA loci, floral-dip and root transformation experiments were carried out in Arabidopsis thaliana with mixtures of A. tumefaciens strains, each harboring one or two different T-DNA vectors. Upon floral-dip transformation, 6-30% of the transformants were co-transformed by multiple T-DNAs originating from different bacteria and 20-36% by different T-DNAs from one strain. However, these co-transformation frequencies were too low to explain the presence of on average 4-6 T-DNA copies in these transformants, suggesting that, upon floral-dip transformation, T-DNA replication frequently occurs before or during integration after the transfer of single T-DNA copies. Upon root transformation, the co-transformation frequencies of T-DNAs originating from different bacteria were similar or slightly higher (between 10 and 60%) than those obtained after floral-dip transformation, whereas the co-transformation frequencies of different T-DNAs from one strain were comparable (24-31%). Root transformants generally harbor only one to three T-DNA copies, and thus co-transformation of different T-DNAs can explain the T-DNA copy number in many transformants, but T-DNA replication is postulated to occur in most multicopy root transformants. In conclusion, the comparable co-transformation frequencies and differences in complexity of the T-DNA loci after floral-dip and root transformations indicate that the T-DNA copy number is highly determined by the transformation-competent target cells.
De Buck
Sylvie
S
Department of Plant Systems Biology, VIB, B-9052 Gent, Belgium.
Podevin
Nancy
N
Nolf
Jonah
J
Jacobs
Anni
A
Depicker
Ann
A
D016960:Agrobacterium tumefaciens / Q000235:genetics*; D017360:Arabidopsis / Q000235:genetics*; D056915:DNA Copy Number Variations; D004269:DNA, Bacterial; D018744:DNA, Plant / Q000235:genetics; D018014:Gene Transfer Techniques; D005822:Genetic Vectors; D018745:Genome, Plant; D030821:Plants, Genetically Modified / Q000235:genetics; D014170:Transformation, Genetic
D004269:DNA, Bacterial; D018744:DNA, Plant; C036483:T-DNA