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First published online October 21, 2005; 10.1104/pp.105.068692

Plant Physiology 139:1338-1349 (2005)
© 2005 American Society of Plant Biologists

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GENETICS, GENOMICS, AND MOLECULAR EVOLUTION

Plant-Derived Transfer DNAs

Caius M. Rommens*, Oleg Bougri, Hua Yan, Jaime M. Humara, Joanna Owen, Kathy Swords and Jingsong Ye

J.R. Simplot Company, Simplot Plant Sciences, Boise, Idaho 83706

The transfer of DNA from Agrobacterium to plant cell nuclei is initiated by a cleavage reaction within the 25-bp right border of Ti plasmids. In an effort to develop all-native DNA transformation vectors, 50 putative right border alternatives were identified in both plant expressed sequence tags and genomic DNA. Efficacy tests in a tobacco (Nicotiana tabacum) model system demonstrated that 14 of these elements displayed at least 50% of the activity of conventional Agrobacterium transfer DNA borders. Four of the most effective plant-derived right border alternatives were found to be associated with intron-exon junctions. Additional elements were embedded within introns, exons, untranslated trailers, and intergenic DNA. Based on the identification of a single right border alternative in Arabidopsis and three in rice (Oryza sativa), the occurrence of this motif was estimated at a frequency of at least 0.8x10–8. Modification of plasmid DNA sequences flanking the alternative borders demonstrated that both upstream and downstream sequences play an important role in initiating DNA transfer. Optimal DNA transfer required the elements to be preceded by pyrimidine residues interspaced by AC-rich trinucleotides. Alteration of this organization lowered transformation frequencies by 46% to 93%. Despite their weaker resemblance with left borders, right border alternatives also functioned effectively in terminating DNA transfer, if both associated with an upstream A[C/T]T[C/G]A[A/T]T[G/T][C/T][G/T][C/G]A[C/T][C/T][A/T] domain and tightly linked cytosine clusters at their junctions with downstream DNA. New insights in border region requirements were used to construct an all-native alfalfa (Medicago sativa) transfer DNA vector that can be used for the production of intragenic plants.

The author responsible for distribution of materials integral to the findings presented in this article in accordance with the policy described in the Instructions for Authors (www.plantphysiol.org) is: Caius M. Rommens (crommens{at}simplot.com).

Article, publication date, and citation information can be found at www.plantphysiol.org/cgi/doi/10.1104/pp.105.068692.

* Corresponding author; e-mail crommens{at}simplot.com; fax 208–327–3212.

Received July 22, 2005; returned for revision July 22, 2005; accepted August 24, 2005.




This article has been cited by other articles:


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H. Yan and C. M. Rommens
Transposition-Based Plant Transformation
Plant Physiology, February 1, 2007; 143(2): 570 - 578.
[Abstract] [Full Text] [PDF]


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