Establishing a foundation for large DNA transfer to artificial minichromosomes and B insert platforms in maize

Abstract

In plants, conventional genetic engineering methods limit the number of available traits that could potentially improve the quality of agriculture. Agrobacterium-mediated transformation and biolistic bombardment are tools used in transferring genes into plant cells, both of which result in random integrations into host genomes. The absence of targeting machinery, together with low DNA carrying capacity on most plasmid vectors, limit researchers to a few genes in a single modification experiment, a process that takes [about]1 year in most plant species. While stacking traits from independent genetic modifications allow for an increase in the number of transgenes in a single plant, recovery of all genes in subsequent generations becomes increasingly difficult due to independent segregation in meiosis. Alternatively, the use of binary bacterial artificial chromosomes (BiBACs), large insert cloning vectors, can maintain and transfer up to 300 kps, but are also subject to random integrations. Therefore, establishment of a BiBAC targeting system would be advantageous for researchers focusing on creating plant lines that contain several genes that work together to express complex traits, such as disease resistance clusters or whole biosynthetic pathways. Additionally, BiBAC targeting to a location outside the native chromosomal sets, such as an artificial minichromosome or B chromosome platform, would enable researchers to stack traits without disrupting endogenous sequences.