New technique for gene transfection using laser irradiation

Y. Shirahata, N. Ohkohchi, H. Itagak, S. Satomi

研究成果: Article査読

63 被引用数 (Scopus)

抄録

Background: We have developed a gene transfection system using laser beams. The principle of this procedure is that a small hole is made in a cell membrane by pulse laser irradiation, and a gene contained in a medium is transferred into the cytoplasm through the hole. This hole disappears immediately with the application of laser irradiation of the appropriate power. Methods: A pulse-wave Nd:YAG laser with a wavelength of 355 nm was used to make a hole in a cell membrane. To trap a cell, a continuous-wave Nd:YAG laser with a wavelength of 1015 nm was used. Plasmids that encode the enhanced green fluorescent protein (EGFP) gene were contained in a medium and transferred to HuH-7 and NIH/3T3 cells with pulse laser irradiation. We evaluated transfection efficiency on the basis of the number of cells that expressed EGFP. Stimulatory protein 2 cells in suspension were fixed using a trapping laser and the neomycin-resistance gene was transfected by pulse laser irradiation. We examined cell proliferation in the selection medium. Results: Cells that expressed EGFP were recognized in the group that was irradiated by pulse laser. No cells expressed EGFP without irradiation. Transfection efficiency was ≈10% at a plasmid concentration of 10.0 μg/mL. At concentrations greater than 20 μg/mL, the transfection rate reached a plateau. We also successfully transfected neomycin-resistance genes to cells floating in suspension after fixation that was achieved with trapping laser irradiation. Conclusions: This method enables us to transfect targeted cells, ie, cells in suspension as well as attached cells, with a simple technique that does not involve harmful vectors. The present method is very useful for gene transfection in cellular biotechnology.

本文言語English
ページ(範囲)184-190
ページ数7
ジャーナルJournal of Investigative Medicine
49
2
DOI
出版ステータスPublished - 2001

ASJC Scopus subject areas

  • 生化学、遺伝学、分子生物学(全般)

フィンガープリント

「New technique for gene transfection using laser irradiation」の研究トピックを掘り下げます。これらがまとまってユニークなフィンガープリントを構成します。

引用スタイル