Targeted gene insertions and dwarfed stature engineering in diploid potato

Dolaptchiev, Yordan Nikolaev (2025) Targeted gene insertions and dwarfed stature engineering in diploid potato. Doctoral thesis, University of East Anglia.

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Abstract

Classical crop breeding in tetraploid potatoes is constrained by difficult genetics, resulting in little genetic gain in yield in recent decades and a widespread susceptibility to important pathogens like Phytophthora infestans. Diploid potato breeding provides a promising alternative, but adoption requires rapid germplasm improvement. While transgenesis can help, the ability to deliver genes at an operator-defined location, here referred to as a knock-in (KI), could allow for more consistent transgene expression and the repeated stacking of transgenes at the same locus. In this thesis, I worked to establish an efficient method for targeted gene delivery in diploid potato. An anthocyanin KI reporter system was established and used to optimise several aspects of the KI approach. This optimised method was used to deliver the P. infestans resistance gene Rpi-vnt1 at the Cai1 locus. Based on this progress, I propose a novel two-phase approach for the targeted iterative insertion of genes via recombination (TIIGER). In the first phase a targeted gene replacement is used to exchange the proximal Cai1 promoter for a cassette with genes of interest, the anthocyanin reporter, and a negative selection marker. In the second phase, the reporter and marker are exchanged for additional genes of interest and the original proximal Cai1 promoter. Here, I demonstrated the first phase and proposed several approaches to the second phase. Lastly, I used gene editing to target the potato DELLA growth regulator gene and generate a dominant mild dwarfism allele which should help improve potatoes’ tolerance to high planting density and improve yields per hectare. A phenotypic series of dwarfs was generated and a promising line bearing a Valine deletion was assessed in a field trial, where it compared favourably to controls. The limitations of these approaches, viable solutions, and future work are also discussed.

Item Type:	Thesis (Doctoral)
Faculty \ School:	Faculty of Science > School of Biological Sciences
Depositing User:	Chris White
Date Deposited:	03 Dec 2025 08:48
Last Modified:	03 Dec 2025 08:48
URI:	https://ueaeprints.uea.ac.uk/id/eprint/101205
DOI:

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