Related Experiment Videos
New technique for gene transfection using laser irradiation.
Y Shirahata1, N Ohkohchi, H Itagak
1Division of Advanced Surgical Science and Technology, Graduate School of Medicine, Tohoku University, Sendai, Japan. sirahata@mail.cc.tohoku.ac.jp
Summary
This study introduces a novel laser-based gene transfection system. The laser creates temporary holes in cell membranes for efficient gene delivery without harmful vectors, proving useful in biotechnology.
Area of Science:
- Biotechnology
- Molecular Biology
- Cell Biology
Background:
- A novel gene transfection system utilizing laser beams has been developed.
- The technique involves creating a transient pore in the cell membrane via pulse laser irradiation to facilitate gene entry.
- The pore reseals immediately after laser application.
Purpose of the Study:
- To develop and evaluate a laser-mediated gene transfection system.
- To assess the efficiency of gene delivery into various cell types.
- To demonstrate the applicability of the system for both attached and suspended cells.
Main Methods:
- Utilized a 355 nm pulse-wave Nd:YAG laser for membrane perforation and a 1015 nm continuous-wave Nd:YAG laser for cell trapping.
- Transfected enhanced green fluorescent protein (EGFP) and neomycin-resistance genes into HuH-7 and NIH/3T3 cells.
- Evaluated transfection efficiency by quantifying EGFP expression and cell proliferation in selection medium.
Main Results:
- Cells successfully expressed EGFP post-transfection, with no expression observed in control groups.
- Achieved a transfection efficiency of approximately 10% at 10.0 microg/mL plasmid concentration, plateauing above 20 microg/mL.
- Demonstrated successful transfection of neomycin-resistance genes into laser-trapped suspension cells.
Conclusions:
- The laser-based system allows for targeted gene transfection of both attached and suspended cells.
- This simple technique avoids the use of harmful vectors, offering a safer alternative.
- The method is highly beneficial for gene transfection applications in cellular biotechnology.