Abstract
The recognition of the T-DNA left border (LB) repeat is affected by its surrounding sequences. Here, the LB regions were further characterized by molecular analysis of transgenic plants, obtained after Agrobacterium tumefaciens-mediated transformation with T-DNA vectors that had been modified in this LB region. At least the 24-bp LB repeat by itself was insufficient to terminate the T-strand synthesis. Addition of the natural inner and/or outer border regions to at least the LB repeat, even when present at a distance, enhanced the correct recognition of the LB repeat, reducing the number of plants containing vector backbone sequences. In tandem occurrence of both the octopine and nopaline LB regions with their repeats terminated the T-strand synthesis most efficiently at the LB, yielding a reproducibly high number of plants containing only the T-DNA. Furthermore, T-strand synthesis did not terminate efficiently at the right border (RB) repeat, which might indicate that signals in the outer RB region inhibit the termination of T-strand synthesis at the RB repeat.
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Acknowledgements
The authors thank Gert Van Der Auwera, Ingrid Peck, and Thierry Aelbrecht for critical reading of the manuscript and many editorial suggestions, Geert Angenon and Willem Broekaert for helpful comments, Vladimir Mironov for generously providing constructs pCDVIB, pCDVIB1, pCDVIB2, and pCDVIB3, Roger Hellens for donating the bar cassette of the pGreen vector, and Martine De Cock for help with the layout. This work was supported by a grant from the European Union BIOTECH program (BIO4-96-0253) and by a STREP project under the 6th framework program of the EU “GENINTEG” (LSHG-CT2003-503303) with additional co-financing from the Flemish Community and a grant from the Special Research Fund of the Ghent University (BOF-01111400). N.P. is indebted to the Institute for the Promotion of Innovation through Science and Technology in Flanders for a predoctoral fellowship.
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Podevin, N., De Buck, S., De Wilde, C. et al. Insights into recognition of the T-DNA border repeats as termination sites for T-strand synthesis by Agrobacterium tumefaciens . Transgenic Res 15, 557–571 (2006). https://doi.org/10.1007/s11248-006-9003-9
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DOI: https://doi.org/10.1007/s11248-006-9003-9