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  • Title: Gene Stacking in Plants Through the Application of Site-Specific Recombination and Nuclease Activity.
    Author: Srivastava V.
    Journal: Methods Mol Biol; 2019; 1864():267-277. PubMed ID: 30415342.
    Abstract:
    Biotechnology methods for inserting genes one by one or as a block of fragment into plant genomes are needed to introduce valuable traits into crop varieties. Insertion of multiple genes into a single site, called as molecular stacking, is important to allow co-inheritance of the genes into the progeny. Generally, two approaches are available for creating gene stacks: nuclease-induced targeted gene integration into native sites and recombinase-mediated gene integration into the engineered sites. The recombinase application is attractive as several recombinases show high efficiency and precision in plant genomes. This chapter describes a gene stacking method based on the use of Cre-lox site-specific recombination system to integrate genes into the engineered sites and nucleases to delete selection genes leading to stacking of traits into a single genomic site. High efficiency and precision, and undetectable off-target effects of Cre-lox in a number of plant species, make it an attractive tool for complex applications such as gene stacking.
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