Question

Write an illustrated account of physical methods of gene transfer.

Answer 1

Physical methods of gene transfer are as follows:

1. Gene gun method
2. Electroporation
3. Lipofection or Liposome mediated gene transfer.
4. Microinjection

1. Gene gun: Gene gun is also known as particle gun, shotgun or microprojectile, etc. By this technique, gene transfer is possible in the intact plant cells. (cells having intact cell wall). This technique of gene transfer was first used by Klein and Co-workers (1987) in onion cells for transferring DNA and viral RNA.
In this process, the minute gold or tungsten particles of 1-3 mm diameter (microparticles) coated with desired DNA are shot (bombarded) in the target cells with the help of a microprojectile.
These desired DNA coated gold or tungsten particles penetrate the cell wall and enter the cell where desired DNA incorporates with the host cell DNA and forms transgenic DNA. By the use of this method, gene transfer has been successfully achieved in wheat, rice, maize, tobacco, and soybean, etc. This technique is being used now in all types of plants worldwide.
2. Electroporation: In this technique of gene transfer, the targeted protoplasts, plant cells, or tissues are subjected to pulse of high voltage. As a result, minute temporary pores are formed in the plasmalemma (plasma membrane).
The desired DNA enters through these minute pores. The target cells or the tissues are kept in a solution containing the desired DNA and then subjected to a high voltage pulse. The DNA from the solution first enters into the cells and then gets incorporated into the genome of the cells. This method is extensively used for transferring genes in the cells of monocotyledonous plants.
3. Liposome mediated gene transfer: In this method of gene transfer, spherical lipid molecules filled with desired DNA and water are used for transferring genes. These DNA containing lipid capsules first stick to the plasmalemma (plasma membrane) and then fuse with it.
The DNA present in these first enters into the cell and then enters the nucleus where it gets incorporated in the genome of the host cell. The liposome directed gene transferring technique also called as lipofection technique is a highly effective technique for transferring genes in bacteria, animal & plant cells.
4. Microinjection: In this method, genes are injected in plant protoplasts or cells with the help of a glass needle of 0.5 - 1.0 mm diameter or micropipette directly in the cytoplasm or nucleus of the protoplasts or cells of plants. This is a suitable method of transferring genes in isolated protoplasts.
Note: Besides the above methods of gene transfer other methods are also employed for transferring genes. These are Ladder dependent gene transfer and Silicon carbide fibre dependent gene transfer.