| Technique |
Procedure |
Most important parameters involved |
Advantages |
Drawbacks |
| Electroporation |
DNA is inserted through pores due to permeabilization of the cell membrane induced by strong electrical pulses. |
Pulse length, energy and duration of the electrical field, extent and duration of membrane permeation, mode and duration of molecular flow, DNA concentration, tolerance of cells to membrane permeation. |
Simple, fast, low cost. |
Low efficiency, requires laborious protocols, and transforms mainly protoplasts. |
| Biolistics |
High density carrier particles covered with genes are accelerated through the cells leaving the DNA inside by an adsorption mechanism. |
Kinetic energy of the bombarding particles, temperature, the amount of cells, their ability to regenerate,susceptibility of the tissue, the number of DNA-coated particles, as well as the amount of DNA that covers each particle. |
Simple, no need to treat the cell wall, allows transformation of different cells, independent of the physiological properties of the cell, allows the use of multiple transgenes. |
High cost, low efficiency Transformation parameters must be optimized to each biological target employed, there is a risk of multiple copies of the introduced genes, DNA and cells can be damage. |
| Agitation with glass beads |
Rapid agitation with glass beads allows the penetration of the plasmid DNA. |
DNA and concentration, sensitivity of cells to membrane permeation, amount of cells and their ability to regenerate. |
Fast, simple, low cost. Does not need sophisticated devices, chemical treatments or enzymatic cocktails. |
Low efficiency because DNA get damaged. |
| Vacuuminfiltration |
Vacuum application generates a negative pressure that increases inter-cell spaces allowing the infiltration of Agrobacterium. |
Duration and intensity of the vacuum, temperature, pH and time of induction of virulence genes. |
Simple, fast, medium efficiency, with low somaclonal variation and many independent cells transformed. |
Some strains of Agrobacterium are unable to infect certain cell types, risk of multiple copies of the introduced genes. |
| Silicon carbide whisker |
Silicon carbide fibers are mixed in a vortex with a suspension of tissue and DNAallowing introduction by abrasion. |
Fiber size, vortex parameters (type, duration and speed of agitation), vessel shape, thickness of the cell wall and cell’s ability to regenerate. |
Simple, fast, low cost and can be used indifferent cell types. |
Very low efficiency. Cells can be damaged affecting regeneration capabilities. Could be hazardous to technicians due to fibers’ inhalation. |
| Lasermicrobeams |
A laser microbeam punctures self-healing holes into the cell wall allowing DNA penetration. |
Laser characteristics to be used as optical tweezers coupled to the appropriate microscope. |
Allows precise and gentle treatment of cells, subcellular structures, and even individual DNA molecules. |
High cost (expensive equipment required), and laborious. |
| Ultrasound |
Introduces DNA molecules into cells via acoustic cavitation that temporarily changes the permeability of the cell membrane. |
Intensity, exposure time, central frequency, type of application (continuous or pulsed), pulse repetition frequency, and duty cycle. |
High efficiency, medium cost and can be used in different cell’s types. |
May damage the cells by breaking their membrane. |
| Shock waves |
Cell permeabilization occurs due to shock wave-induced cavitation. |
Frequency, energy, voltage, shock wave profile and number of shock waves. |
Fast, easy to perform, reproducible with high efficiency, no need of enzymatic cocktails, can be used to transform several cell types. |
Shock wave generators for this purpose are not on the market yet and experimental devices are relatively expensive. |
