White LED intensities during co-cultivation affect the Agrobacterium-mediated soybean (Glycine max) transformation using mature half seeds as explants

Abstract

The transition of light fixture from fluorescent light to light-emitting diodes (LEDs) in growth chambers prompts a reevaluation of current practices in plant biotechnology. Agrobacterium-mediated transformation is crucial for genetic engineering and genome editing in soybean (Glycine max). The critical co-cultivation step of soybean transformation occurs under light condition. Current protocols for co-cultivation in soybean transformation lack a standard for light intensity. In the present study, the objective is to investigate the effect of light intensity during co-cultivation on soybean transformation efficiency. Five light intensities were implemented during five days of co-cultivation: 50, 100, 150, 190 μmol∙m−2∙s−1 of white LEDs in addition to 100 μmol∙m−2∙s−1 of fluorescent light. After co-cultivation, all the explants underwent shoot induction and elongation with selection pressure, rooting and acclimation under uniform condition. The experiment was conducted with two selectable markers, hppdPf-4Pa and bar, separately, investigating whether the potential light effects vary due to the marker-associated pathways. The positive PCR analysis of rooted in vitro plants suggested successful transformation events achieved under both selectable markers across all light treatments ranging from 2.4% to 6.9%. Increasing LED light intensity during co-cultivation resulted in different transformation efficiencies between the two selectable markers. Results indicated that increasing the light intensity during co-cultivation led to a linear increase in transformation efficiency when shoot regeneration was under 4-Hydroxyphenylpyruvate dioxygenase (HPPD) inhibitor selection. No difference in transformation efficiency was detected among the treatments under glufosinate selection. Furthermore, when selection occurred with HPPD inhibitor, variation of transformation efficiency was also observed between fluorescent light and white LED at 100 μmol∙m−2∙s−1. The results highlight the significance and potential applications of investigating the impact of light on transformation efficiency.