Why Transfect Cells?
Scientists transfect cells to understand gene function and produce specific proteins. Introducing foreign genetic material allows researchers to study how individual genes influence cellular processes like growth, differentiation, or disease development. This can involve overexpressing a protein to observe its effects or reducing gene expression to understand its role.
Transfection is also used to produce recombinant proteins, engineered to be expressed in cells different from their natural source. These proteins serve as research tools, therapeutic agents like insulin or antibodies, or components for vaccine development. The technique also aids in creating cellular models of human diseases, allowing scientists to study disease mechanisms and test potential drug candidates. Ultimately, the ability to manipulate gene expression within cells provides a powerful approach for both fundamental biological discovery and the development of new biotechnologies.
How Transfection Works
Introducing foreign genetic material into eukaryotic cells requires overcoming the cell’s natural barriers, particularly the cell membrane, which typically restricts the entry of large molecules like DNA and RNA. Specialized methods facilitate this uptake, generally categorized into chemical, physical, and sometimes biological approaches. Each method aims to temporarily disrupt the cell membrane or create pathways for the genetic material to enter the cytoplasm.
Chemical methods utilize positively charged compounds like lipids or polymers that form complexes with the genetic material. These complexes interact with the negatively charged cell membrane, leading to their internalization. For instance, lipofection uses liposomes, lipid vesicles that encapsulate DNA or RNA and fuse with the cell membrane, releasing contents inside. These agents help shield the genetic material from degradation.
Physical methods directly create transient pores or openings in the cell membrane. Electroporation applies short, high-voltage electrical pulses to cells, temporarily increasing membrane permeability. This allows genetic material to diffuse into the cell before pores reseal. Another physical method, microinjection, uses a fine needle to directly inject genetic material into individual cells, offering precise delivery. While this article focuses on non-viral transfection, it’s worth noting that modified viruses are also used to deliver genetic material, a process often called transduction.
Different Types of Transfection
Transfection can result in two primary outcomes based on how the introduced genetic material behaves within the host cell: transient or stable expression. The choice between these types depends on experimental goals and the desired duration of gene expression.
Transient transfection occurs when the introduced genetic material, typically DNA, remains in the cell’s cytoplasm and does not integrate into the host cell’s chromosomes. This genetic material is expressed for a limited period, usually a few days, before being degraded by cellular enzymes or diluted by cell division. Transient transfection is often used for quick experiments, such as testing gene function or producing small quantities of protein, as it offers rapid results without lengthy selection processes.
In contrast, stable transfection involves the integration of the introduced genetic material into the host cell’s genome. This means the new gene becomes a permanent part of the cell’s genetic makeup, passed to all subsequent daughter cells. To achieve stable transfection, cells are cultured under selective conditions, often using an antibiotic resistance gene carried on the same genetic construct, allowing only cells that have successfully integrated the foreign DNA to survive. Stable transfection is employed when long-term, continuous expression is required, such as in creating cell lines or for gene therapy applications.