Introduction of protein into plant cells using atmospheric temperature controllable plasma with accuracy within 3.8 K

In recent years, genome editing has attracted attention as a prominent method to improve crop varieties. However, introducing genome editing enzymes into plant cells remains a challenge due to the barrier such as cell walls and cuticles. In our group, atmospheric plasma has considered as a solution. To irradiate plasma to heat-sensitive plants without damaging, we developed the world’s first atmospheric temperature controllable plasma system which can control gas temperatures from 3 to 10°C. Using plasma controlled at 20 to 30°C, we successfully introduced genome editing enzymes into tobacco cells. During this research, it was suggested that plasma gas temperature may affect protein introduction. However, this system showed deviations of about ±5°C from setting temperature, making it difficult to evaluate this effect accurately.

In this study, we developed a new plasma system which can control plasma gas temperature with ±1.9 K accuracy. Using this system, we irradiated plasma precisely controlled at different temperatures to onion epidermis and evaluated the amount of protein introduced. N₂ plasma controlled from near 0 to 100°C was irradiated. The plasma irradiated onion tissue was incubated overnight with a solution containing red fluorescent protein (His-TagRFP).

RFP fluorescence in cells was observed under temperature conditions from 10 to 25°C indicating successful protein introduction. Comparisons of introduced protein amount at different plasma gas temperature will be reported.