Location: Crop Improvement and Genetics ResearchTitle: Recombinase technology for precise genome engineering Author
|Thomson, James - Jim|
Submitted to: Book Chapter
Publication Type: Book / Chapter
Publication Acceptance Date: 3/3/2015
Publication Date: 5/25/2015
Citation: Thomson, J.G., Blechl, A.E. 2015. Recombinase technology for precise genome engineering. In: Zhang, F., Puchta, H., Thomson, J.G., editors. Advances in new technology for targeted modification of plant genomes. New York, NY: Springer. p. 113-144. Interpretive Summary: This article summarizes the impact that site-specific recombination-based technologies are having on the field of genomic engineering, and how these methods will change genomic engineering in the future. This technology enables the removal of extraneous DNA such as selectable markers, (i.e. antibiotic resistance genes) from the genome as well as speeds the transition from laboratory manipulation to field production; an important benefit for both the industry and general public. From an application point of view, the paper discusses the number and type of recombinases available along with the innovative strategies being developed. It offers a multitude of recombinase applications for genome manipulation, with everything from single copy high throughput targeted integration, to sequential gene stacking, to complete transgene removal from the pollen and/or seed.
Technical Abstract: The use of recombinases for genomic engineering is no longer a new technology. In fact this technology has entered its third decade since the initial discovery that recombinases function in heterologous systems. The random insertion of a transgene into a plant genome by traditional methods generates unpredictable expression patterns. This feature of transgenesis makes screening for functional lines with predictable expression labor intensive and time consuming. Furthermore, an antibiotic resistance gene is often left in the final product and the potential escape of such resistance markers into the environment and their potential consumption raises consumer concern. The use of site-specific recombination technology in plant genome manipulation has been demonstrated to effectively resolve complex transgene insertions to single copy, remove unwanted DNA, and precisely insert DNA into known genomic target sites. Recombinases have also been demonstrated capable of site-specific recombination within non-nuclear targets, such as the plastid genome of tobacco. Here, we review multiple uses of site-specific recombination and their application toward plant genomic engineering. We also provide alternative strategies for the combined use of multiple site-specific recombinase systems for genome engineering to precisely insert transgenes into a pre-determined locus, and removal of unwanted selectable marker genes