Acidic pH Condition Enhances Femtosecond Laser Photoinjection of Plasmid DNA into Intact Tobacco BY‑2

In plant biology, targeted photoinjection is a promising approach to understand the role of individual cells in the context of the overall structure. In particular, photomechanical poration by a femtosecond laser combined with partial degradation of cell wall structure by an enzyme treatment has been shown to be a great advantage for the injection of macromolecules. To extend the femtosecond laser technique to plasmid DNA, the effect of the electrostatic interaction between plasmid DNA and the cell wall during the photoinjection processes was investigated in this work. An imaging assay utilizing fluorescence-labeled plasmid DNA revealed that plasmid DNA molecules were accumulated into the inner cell wall under conventional cell culture conditions. It is likely that the electrical potential of the cell wall and the plasmid DNA were modified under acidic pH conditions. In addition, the cell wall was softened according to the acid growth theory. These combined effects lead to efficient photoinjection of plasmid DNA through the pores created by femtosecond laser irradiation.