Uses of transfection

Why do we use transfection? There are several uses of transfection in molecular biology. 

It is used for a range of molecular biology applications. 

It is widely used for creating knockout mice. 

Gene therapy: The idea behind this is that a defective gene is replaced with a function gene or even down regulating an over expressed gene or an oncogene. 

Transfection methods

There are 3 most widely used transfection methods. 

Transfection methods can be decided into 3 categories 

1. viral and non-viral methods. Although the uses of viruses are not really transfection but transduction, it is used and is very efficient. 

2. of the most widely used viruses are lentivirus and adenovirus. 

3. chemical methods are the most widely used method for transecting cells. 

Optimisation and efficiency 

Most of the widely used transfection is though the use of chemicals. It is important to optimize transfection conditions for optimal transfection efficiencies. 

Time of incubation is important as cells may loose viability. You will have monitor cells morphology every few hours especially serum is not used.

Transfection methods 

Calcium precipitate and DAE Dextran: Formation of calcium precipitates that produce a net positive charge and associate with the negatively charged cell membrane and internalise through endocytosis. 

Liposome-mediated transfection (lipofection Transfection Reagent results in stable and transient transfections with low cytotoxicity and consistent high transfection efficiencies

(a 10-fold increase over calcium phosphate/DEAE mediated transfection in many cell lines).

Electroporation: This method takes advantage of the fluid nature if the lipid bilayer of the cell membrane and the ability to reassemble after being disturbed. 

The technique requires fine-tuning and optimization for duration and strength of the pulse for each type of cell used. A critical balance must be achieved between conditions that allow efficient delivery and conditions that kill cells.

External current (300-400 mV) is used to increase conductivity and permeability of the cell membrane. The current rapidly changes the morphology of the lipid bilayer. This created something called a pre-pore allowing insertion of DNA in the cells. It is very useful in introducing genes and creating knockout mice.