Excision of DNA fragments with the piggyBac system in Chrysanthemum morifolium

Mitsuko Kishi-Kaboshi; Ayako Nishizawa-Yokoi; Ichiro Mitsuhara; Seiichi Toki; Katsutomo Sasaki

doi:10.5511/plantbiotechnology.23.0324a

Excision of DNA fragments with the piggyBac system in Chrysanthemum morifolium

Plant Biotechnol (Tokyo). 2023 Jun 25;40(2):157-165. doi: 10.5511/plantbiotechnology.23.0324a.

Authors

Mitsuko Kishi-Kaboshi¹, Ayako Nishizawa-Yokoi², Ichiro Mitsuhara³, Seiichi Toki^{2

4}, Katsutomo Sasaki¹

Affiliations

¹ Institute of Vegetable and Floriculture Science, National Agriculture and Food Research Organization, 2-1 Fujimoto, Tsukuba, Ibaraki 305-0852, Japan.
² Institute of Agrobiological Sciences, National Agriculture and Food Research Organization, 3-1-3 Kannondai, Tsukuba, Ibaraki 305-8604, Japan.
³ Institute of Agrobiological Sciences, National Agriculture and Food Research Organization, 2-1-2 Kannondai, Tsukuba, Ibaraki 305-8602, Japan.
⁴ Laboratory of Plant Genome Engineering, Department of Plant Life Science, Faculty of Agriculture, Ryukoku University, 1-5 Yokotani, Seta Oe-cho, Otsu, Shiga 520-2194, Japan.

PMID: 38250294
PMCID: PMC10797517
DOI: 10.5511/plantbiotechnology.23.0324a

Abstract

Chrysanthemum morifolium is one of the most popular ornamental plants in the world. However, as C. morifolium is a segmental hexaploid, self-incompatible, and has a sizable heterologous genome, it is difficult to modify its trait systematically. Genome editing technology is one of the attractive methods for modifying traits systematically. For the commercial use of genetically modified C. morifolium, rigorous stabilization of its quality is essential. This trait stability can be achieved by avoiding further genome modification after suitable trait modification by genome editing. Since C. morifolium is a vegetatively propagated plant, an approach for removing genome editing tools is required. In this study, we attempted to use the piggyBac transposon system to remove specific DNA sequences from the C. morifolium genome. Using the luminescence as a visible marker, we demonstrated that inoculation of Agrobacterium harboring hyperactive piggyBac transposase removes inserted 2.6 kb DNA, which harbors piggyBac recognition sequences, from the modified Eluc sequence.

Keywords: Chrysanthemum morifolium; DNA excision; genome engineering; piggyBac; transposon.