Biolistics
Introduction
Biolistics, also known as ballistic particle delivery, is a technique widely used in biotechnology that involves the delivery of DNA into cells using high velocity microprojectiles. The term "biolistics" is a portmanteau of "biological ballistics". This method is often employed in plant transformation but has also been adapted for use in animal cells, including mammalian cells.
History and Development
The concept of biolistics was first introduced in the 1980s by John C. Sanford and his colleagues at Cornell University. The initial method was crude and involved the use of a .22 caliber gunpowder charge to propel the DNA-coated microprojectiles. Over time, the technique has been refined and commercialized, with the most common device now being the gene gun.
Principle and Procedure
The principle behind biolistics is relatively straightforward. DNA is precipitated onto small metal particles, usually gold or tungsten. These DNA-coated particles are then accelerated to high speeds, typically using a burst of high pressure helium gas. The particles penetrate the cell wall and membrane, delivering the DNA directly into the cell's cytoplasm or nucleus.
The procedure of biolistics involves several steps. First, the DNA is precipitated onto the metal particles. This is usually done by adding the DNA to a solution of the metal particles, followed by the addition of a high concentration of calcium chloride and a brief period of heat shock. The DNA-coated particles are then washed and resuspended in ethanol.
The next step is the actual delivery of the DNA. The DNA-coated particles are loaded into the gene gun and the device is fired at the target cells. The high velocity of the particles allows them to penetrate the cell wall and membrane, delivering the DNA into the cell.
Applications
Biolistics has a wide range of applications in various fields of biotechnology. It is most commonly used for plant transformation, where it has been used to introduce new genes into a wide variety of plant species. This has allowed for the development of genetically modified crops with improved traits such as resistance to pests or diseases, or enhanced nutritional content.
In addition to plant transformation, biolistics has also been used for the transformation of animal cells. This includes the transformation of mammalian cells, where it has been used for the introduction of genes for the production of recombinant proteins. It has also been used for the transformation of insect cells, for the production of insect-resistant crops.
Biolistics has also been used in the field of vaccinology, for the delivery of DNA vaccines. This involves the introduction of a gene encoding an antigen into the cells of the recipient, which then produce the antigen and stimulate an immune response.
Advantages and Limitations
One of the main advantages of biolistics is that it can be used to transform a wide variety of cell types, including those that are difficult to transform by other methods. It is also a relatively simple and straightforward procedure, which does not require the use of complex equipment or reagents.
However, there are also several limitations to the use of biolistics. One of the main limitations is that it can cause damage to the cells, due to the high velocity of the particles. This can result in a low efficiency of transformation. Another limitation is that it can be difficult to control the number of copies of the gene that are introduced into the cells, which can result in variable expression levels.
Conclusion
In conclusion, biolistics is a powerful tool in biotechnology, with a wide range of applications. Despite its limitations, it remains a commonly used method for the transformation of both plant and animal cells.