Direct protein introduction into plant cells using a multi-gas plasma jet.
Protein introduction into cells is more difficult in plants than in mammalian cells, although it was reported that protein introduction was successful in shoot apical meristem and leaves only together with a cell-penetrating peptide. In this study, we tried to introduce superfolder green fluorescent protein (sGFP)-fused to adenylate cyclase as a reporter protein without a cell-penetrating peptide into the cells of tobacco leaves by treatment with atmospheric non-thermal plasmas. For this purpose, CO2 or N2 plasma was generated using a multi-gas plasma jet. Confocal microscopy indicated that sGFP signals were observed inside of leaf cells after treatment with CO2 or N2 plasma without substantial damage. In addition, the amount of cyclic adenosine monophosphate (cAMP) formed by the catalytic enzyme adenylate cyclase, which requires cellular calmodulin for its activity, was significantly increased in leaves treated with CO2 or N2 plasma, also indicating the introduction of sGFP-fused adenylate cyclase into the cells. These results suggested that treatment with CO2 or N2 plasma could be a useful technique for protein introduction into plant tissues.
Institute of Agrobiological Sciences, NARO, Kannondai, Tsukuba, Ibaraki, Japan.
Yuki
Y
Yanagawa
FIRST, Institute of Innovative Research, Tokyo Institute of Technology, Nagatsuta, Midori-ku, Yokohama, Japan.
Hiroaki
H
Kawano
FIRST, Institute of Innovative Research, Tokyo Institute of Technology, Nagatsuta, Midori-ku, Yokohama, Japan.
Tomohiro
T
Kobayashi
FIRST, Institute of Innovative Research, Tokyo Institute of Technology, Nagatsuta, Midori-ku, Yokohama, Japan.
Hidekazu
H
Miyahara
FIRST, Institute of Innovative Research, Tokyo Institute of Technology, Nagatsuta, Midori-ku, Yokohama, Japan.
Akitoshi
A
Okino
Institute of Agrobiological Sciences, NARO, Kannondai, Tsukuba, Ibaraki, Japan.
Ichiro
I
Mitsuhara
D000262:Adenylyl Cyclases / Q000235:genetics / Q000378:metabolism*; D002245:Carbon Dioxide / Q000737:chemistry; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism*; D018519:Meristem / Q000378:metabolism*; D009584:Nitrogen / Q000737:chemistry; D018515:Plant Leaves / Q000378:metabolism*; D058626:Plasma Gases; D011312:Pressure; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism
D058626:Plasma Gases; D011994:Recombinant Proteins; D002245:Carbon Dioxide; D049452:Green Fluorescent Proteins; D000262:Adenylyl Cyclases; D009584:Nitrogen