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. 2022 Dec 6:13:1085694.
doi: 10.3389/fgene.2022.1085694. eCollection 2022.

Establishment of an efficient regeneration and Agrobacterium transformation system in mature embryos of calla lily (Zantedeschia spp.)

Xuan Sun  1   2 , Yi Wang  2   3 , Tuo Yang  3 , Xue Wang  2 , Huanxiao Wang  1   2 , Di Wang  1   2 , Hongyan Liu  1   2 , Xian Wang  2 , Guojun Zhang  1   4 , Zunzheng Wei  2
Affiliations

Establishment of an efficient regeneration and Agrobacterium transformation system in mature embryos of calla lily (Zantedeschia spp.)

Xuan Sun et al. Front Genet. .

Abstract

Calla lily (Zantedeschia spp.) have great aesthetic value due to their spathe-like appearance and richness of coloration. However, embryonic callus regeneration is absent from its current regeneration mechanism. As a result, constructing an adequate and stable genetic transformation system is hampered, severely hindering breeding efforts. In this research, the callus induction effectiveness of calla lily seed embryos of various maturities was evaluated. The findings indicated that mature seed embryos were more suitable for in vitro regeneration. Using orthogonal design experiments, the primary elements influencing in vitro regeneration, such as plant growth regulators, genotypes, and nanoscale materials, which was emergent uses for in vitro regeneration, were investigated. The findings indicated that MS supplemented with 6-BA 2 mg/L and NAA 0.1 mg/L was the optimal medium for callus induction (CIM); the germination medium (GM) was MS supplemented with 6-BA 2 mg/L NAA 0.2 mg/L and 1 mg/L CNTs, and the rooting medium (RM) was MS supplemented with 6-BA 2 mg/L NAA 0.7 mg/L and 2 mg/L CNTs. This allowed us to verify, in principle, that the Agrobacterium tumefaciens-mediated genetic transformation system operates under optimal circumstances using the GUS reporter gene. Here, we developed a seed embryo-based genetic transformation regeneration system, which set the stage for future attempts to create new calla lily varieties.

Keywords: CNTs; calla lily; genetic transformation; regeneration system; seed embryo.

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Conflict of interest statement

The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.

Figures

FIGURE 1
FIGURE 1
Embryonic callus induced from hybrid seeds embryo of cv. “Black Magic” × “Liming” at different maturity. (A) Induction of seed embryos at different stages and callus at 21 days. Scale bars, 500 μm (B) Callus induction rates of seed embryos of different maturity levels. (C) Paraffin section observation of embryogenic callus.
FIGURE 2
FIGURE 2
Callus induction in hybrid seeds embryo of calla lily by different orthogonal design.
FIGURE 3
FIGURE 3
Effects of different factors on callus induction, germination, and rooting of hybrid seed embryos of calla lily. (A) Effects of different factors on callus induction. (B) Effects of different concentration of NAA, 6-BA and CNTs on callus germination. (C) Effects of different concentration of NAA, 6-BA and CNTs on callus rooting.
FIGURE 4
FIGURE 4
Somatic embryo induction and genetic transformation of calla lily. (A) A1: Seed embryo inoculation; A2: Seed embryo germination; A3: Seed embryo produces callus; A4: Callus propagation; A5: Callus germination; A6: Elongation; A7: Take root culture; A8: Intact plant; A9: Infect; A10: Co-culture; A11: GUS staining control group; A12: GUS staining test group; Scale bars, 500 μm. (B) Identification of transformation effect by PCR of the GUS fragment, and M represents the DNA marker (C) Identification of GUS expression in transformed callus.
FIGURE 5
FIGURE 5
Schematic diagram of seed embryo regeneration system of calla lily.
See this image and copyright information in PMC

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