Development of an Improved Construct for Spectinomycin Selection in Plant Transformation

Authors: Mc Cue, Kent; SHAO, MIN - Oak Ridge Institute For Science And Education (ORISE); Huynh, Jennie; MINH, TAI - Alpine Bio; CHAN, MANDY - Alpine Bio; MOY, YORK - Alpine Bio; Thomson, James

Submitted to: BMC Research Notes
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 3/7/2025
Publication Date: 3/22/2025
Citation: Mc Cue, K.F., Shao, M., Huynh, J.N., Minh, T., Chan, M., Moy, Y., Thomson, J.G. 2025. Development of an Improved Construct for Spectinomycin Selection in Plant Transformation. BMC Research Notes. 18(1), 122 (2025). https://doi.org/10.1186/s13104-025-07178-3.
DOI: https://doi.org/10.1186/s13104-025-07178-3

Interpretive Summary: In this study we modify the spectinomycin selection marker gene for improved transformation efficiency in multiple plant species. This was accomplished by fusing the sul1 ORF with a truncated stroma targeting domain to the aadA marker gene. The fusion gene cassette, (termed SuSp), was evaluated in Arabidopsis thaliana, Carrizo citrange trifoliate (citrus rootstock), Solanum tuberosum (potato), and Glycine max (soybean) for transformation efficiency as compared to the commonly used spectinomycin (SSp) and kanamycin (nptII) selection marker genes. Results show the enhanced SuSp cassette is superior to the standard spectinomycin and equal to kanamycin selection in Arabidopsis, citrus and potato.

Technical Abstract: The efficient selection of transgenic plants is essential in plant biotechnology, and the development of effective selectable marker systems plays a crucial role in this process, especially in context of multiple rounds of transformation and gene stacking which requires multiple selectable markers. This study aimed to develop and evaluate an enhanced spectinomycin resistance construct to provide robust spectinomycin selection for transformation events. The new construct was tested across multiple plant species and compared to the standard spectinomycin selectable marker gene currently used for soybean transformation and the nptII kanamycin resistance gene. In Arabidopsis thaliana, potato and citrus the enhanced spectinomycin marker provided transformation efficiency approximately equal to kanamycin and considerably better than that observed for the standard spectinomycin marker. In soybean, the enhanced spectinomycin resistance marker preformed equally well to the standard spectinomycin gene when tested. These results demonstrate that the enhanced spectinomycin construct provides strong resistance, comparable to kanamycin in multiple species, offering a superior tool for transgenic plant selection.