What are the Common Methods of Gene Transfer into Host Cells?

The recombinant vector carrying foreign DNA needs to be transferred into the suitable host cells. Several methods have been developed for introduction of recombinant DNA molecule into host cells.

According to types of vectors and host cells, the methods are adopted. Some of the methods of gene transfer into host cells are briefly discussed below:

1. Transformation :

Introduction of rDNA molecules into a living cell is called transformation. The DNA molecule comes in the contact of cell surface. Then it is taken up by the host cells.

However, in nature the frequency of transformation of many cells (e.g. yeast and mammalian cells) is very less. Secondly, all the time host cells do not undergo transformation. Because they are not prepared for it.

When there develops competence factors in the cells, transformation phenomenon occurs. There are some factors which affect transformation such as concentration of foreign DNA molecules, host’s cell density, temperature, etc.

A temperature sock of either low temperature (0-5°C) or high temperature (37- 45°C) is required for transformation. Mandel and Higa (1970) reported that E. coli cells become competent to uptake DNA by treating cells with ice cold CaCl₂ and exposing the cells to 42°C for about 90 seconds.

2. Transfection :

Transfection is the transfer of foreign DNA into cultured host cells mediated through chemicals.

The charged chemical substances such as cationic liposomes, calcium phosphate of DEAE dextran are taken and mixed with DNA molecules. The recipient host cells are overlayed by this mixture. Consequently the foreign DNA is taken up by the host cells.

3. Electro oration (Electric Field-mediated Membrane Permeation) :

In electroporation an electric current at high voltage (about 350 V) is applied in a solution containing foreign DNA and fragile host cells. This creates transient microscopic pores in cell membrane of naked protoplasts.

Consequently foreign DNA enters into the protoplast through these pores. The transformed protoplasts are cultured in vitro which regenerate respective cell walls.

4. Microinjection :

In this technique foreign DNA is directly and forcibly injected into the nucleus of animal and plant cells through a glass micropipette containing very fine tip of about 0.5 mm diameter.

It resembles with injection needle. In 1982, for the first time Rubin and Spradling introduced Drosophila gene into P-element and microinjected into embryo. In 1982, R.D. Palmiter and R.L.

Brinter introduced rabbit recombinant pBR322 containing a growth hormone gene into mature embryo of mouse in vitro. The microinjected embryos were transferred into uterus of mice which gave birth to transgenic mice.

5. Particle Bombardment Gun (Biolistics) :

This technique was developed by Stanford in 1987. In this method macroscopic gold or tungusten particles are coated with desired DNA. A plastic micro-carrier containing DNA coated gold/tungusten particles are placed near rupture disc.

The particles are bombarded onto target cells by the bombardment apparatus. Consequently foreign DNA is forcibly delivered into the host cells.

6. Agrobacterium-mediated Gene Transfer :

As discussed earlier that foreign DNA is inserted into Ti-plasmid of Agrobacterium tumifaciens. The recombinant plasmid is transferred into A. tumifaciens cells. The genetically modified A. tumifaciens cells are co-cultured with plant cells in vitro.

Genetically modified A. tumifaciens infects the cultured plant cell and delivers foreign DNA containing plasmid into the infected cell.

A similar technique is also used to transfer foreign gene into root cells by using genetically modified Agrobacterium rhizogenes cells.