T-DNA integration: a mode of illegitimate recombination in plants

Abstract

Transferred DNA (T-DNA) insertions of Agrobacterium gene fusion vectors and corresponding insertional target sites were isolated from transgenic and wild type Arabidopsis thaliana plants. Nucleotide sequence comparison of wild type and T-DNA-tagged genomic loci showed that T-DNA integration resulted in target site deletions of 29-73 bp. In those cases where integrated T-DNA segments turned out to be smaller than canonical ones, the break-points of target deletions and T-DNA insertions overlapped and consisted of 5-7 identical nucleotides. Formation of precise junctions at the right T-DNA border, and DNA sequence homology between the left termini of T-DNA segments and break-points of target deletions were observed in those cases where full-length canonical T-DNA inserts were very precisely replacing plant target DNA sequences. Aberrant junctions were observed in those transformants where termini of T-DNA segments showed no homology to break-points of target sequence deletions. Homology between short segments within target sites and T-DNA, as well as conversion and duplication of DNA sequences at junctions, suggests that T-DNA integration results from illegitimate recombination. The data suggest that while the left T-DNA terminus and both target termini participate in partial pairing and DNA repair, the right T-DNA terminus plays an essential role in the recognition of the target and in the formation of a primary synapsis during integration.