Transfection: A Core Science Technique
- NanoEntek
- Aug 26
- 3 min read
Cell transfection is the process of deliberately introducing foreign genetic materials (like DNA, RNA) into eukaryotic cells. Think of it as delivering a tiny instruction manual or a new tool into a cell, telling it to do something or to produce a specific protein.
Unlike bacterial transformation, which involves introducing DNA into prokaryotic cells, transfection is specifically used for eukaryotic cells, which include those with a nucleus, such as human, animal, or plant cells.
What is the Purpose of Transfection?
The purpose is to modify gene expression, either by adding new genetic materials (gene overexpression), silencing endogenous genes (RNA interference), or editing genomes (CRISPR-Cas9).
Where is Transfection Used the Most
1. Drug screening and discovery
Transfected cells can be used to screen gene targets or validate therapeutic candidates by expressing specific receptors or disease markers.
2. Gene therapy
Introducing healthy genes into defective cells from patients leads to the expectation of genetic disorder treatment.
3. Protein production
Proteins such as insulin, monoclonal antibodies, enzymes, etc., are produced for research purposes by transfecting cells with the corresponding genes.
4. CRISPR-Cas9 gene editing
Transfection is the primary method to deliver CRISPR-Cas9 components (guide RNA and Cas9 enzyme) into cells to precisely edit their genomes for research or therapeutic purposes.
Main Purposes
The main purpose of cell transfection is to alter the genetic makeup and the behavior or characteristics of cells for scientific investigation or therapeutic benefit.
Gene expression
- To express a protein of interest or to understand gene function, or to introduce plasmid DNA.
Gene silencing
- To knock down (or turn off) specific genes using RNA interference (RNAi) to study their role in pathways or diseases.
Reporter gene assay
- To track gene expression, protein localization, cellular processes, and responses by introducing reporter genes such as GFP or luciferase.
Methodology: How is it done?
Chemical method
- By using chemical reagents, it forms complexes with the nucleic acid, which then facilitates its entry into the cell.
Pros | Easy and cost-effective; stable |
Cons | Possibility of toxicity; pH sensitive |
a) Lipofection (lipid-based transfection)
One of the commonly used methods, which uses lipid-based reagents to form complexes with nucleic acids, and then is taken up by cells through endocytosis. Cationic (positively charged) lipids form liposomes (tiny lipid spheres) that encapsulate the negatively charged DNA or RNA. These liposome-nucleic acid complexes then fuse with the negatively charged cell membrane, allowing the genetic material to enter the cell.
Examples: Lipofectamine, fugene
b) Calcium phosphate transfection
DNA is mixed with calcium chloride and phosphate buffer, forming a precipitate. These DNA-calcium phosphate precipitates are then taken up by the cells through endocytosis.
Physical method
- Physically creates temporary pores or openings in the cell membrane, allowing genetic materials to enter.
Pros | Suitable for difficult cells and large molecules. |
Cons | May require specialized equipment; can damage cells. |
a) Electroporation
Cells are subjected to a brief, high-voltage electrical pulse. This pulse creates transient pores in the cell membrane, through which the nucleic acid can diffuse into the cytoplasm.
b) Microinjection
Nucleic acid is injected into the nucleus or cytoplasm using fine needles. This method is precise and efficient but requires a highly trained person, making it suitable for studies on a small number of cells.
Viral method (transduction)
- While technically called transduction rather than transfection, this method uses modified viruses as vectors to deliver genetic material into cells. Viruses naturally evolved efficient mechanisms to infect cells and deliver their genetic cargo.
Pros | High efficiency, especially in primary cells and in vivo applications. |
Cons | More complex and costly; biosafety considerations required. |
a) Retroviral and Lentiviral Vectors
These viruses integrate their genetic material into the host cell's genome, leading to stable, long-term gene expression. They are excellent for gene therapy applications and creating stable cell lines.
b) Adenoviral Vectors
These viruses deliver DNA into the cell nucleus but typically do not integrate into the host genome, leading to transient expression. They are often used for high-level, short-term protein production.
Conclusion
Cell transfection continues to evolve with the development of new reagents, more efficient devices, and novel viral vectors. As our understanding of cellular processes deepens and gene-editing technologies advance, transfection will remain a vital tool, pushing the boundaries of what is possible in medicine, biotechnology, and fundamental biological research.
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