Recombinase-mediated integration of a multigene cassette in rice leads to stable expression and inheritance of the stacked locus

BioRxiv : the Preprint Server for Biology
B. Pathak, Vibha Srivastava


Efficient methods for multigene transformation are important for developing novel crop varieties. Methods based on random integrations of multiple genes have been successfully used for metabolic engineering in plants. However, efficiency of co-integration and co-expression of the genes could present a bottleneck. Recombinase-mediated integration into the engineered target sites is arguably a more efficient method of targeted integration that leads to the generation of stable transgenic lines at a high rate. This method has the potential to streamline multigene transformation for metabolic engineering and trait stacking in plants. Therefore, empirical testing of transgene(s) stability from the multigene site-specific integration locus is needed. Here, the recombinase technology based on Cre-lox recombination was evaluated for developing multigenic lines consisting of a 5 gene cassette harboring constitutively-expressed and inducible genes. Targeted integration of this cassette in the rice genome generated a precise full-length integration of the cassette into the target site at a high rate, and the resulting multigenic lines expressed each gene reliably as defined by their promoter activity. The stable constitutive or inducible ...Continue Reading

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