The first generation gene gun is a desktop gene gun, and the gunpowder desktop gene gun is the most primitive gene gun. The earliest gene gun was a new gene transfer method developed by Sanford of Cornell University in the United States in 1987 in cooperation with engineering experts Wolff and Karen of the school. Once this method was invented, it made its mark in academic circles. Klein et al. transformed onion epidermal cells with gene gun for the first time in 1987, and achieved success.
Gene gun has developed rapidly since the birth of 1987. Since 1988, Cornell University in the United States has successively applied for three patents of gene gun technology (EP 033 1 855A2, 1988: U.S. Patent No.4,945,050 July 31,1990: U.S. PatentNo.. During the period of 1987- 1990, various gene guns, such as high voltage discharge and compressed gas drive, appeared one after another, all of which were constantly improved and developed through repeated practice. 1988, McCabe wrapped the target gene with tungsten powder and bombarded the meristem of soybean shoot tip. As a result, about 2% of the tissues obtained regenerated plants through organogenesis, and foreign genes were detected in the offspring. 1989 pneumatic gene gun successfully transformed tobacco and other plants and obtained instantaneous expression. Compared with other plants, the development of transgenic technology in barley is relatively slow. Until 1989, Kartha and others cultured barley cells and tissues and successfully detected the transient expression of the reporter gene.
1990, DuPont Company of the United States launched the first commercial gene gun PDS- 1000 system. This instrument is a "Biologics" desktop gene gun, and the related technology was purchased from a small Biologics company (the person in charge is Cornell University). According to W.Haeussler, vice chairman of Cornell Research Foundation and head of university patent and technology market, the technology transferred to DuPont was the biggest transaction invented by Cornell at that time, and a total of 2.28 million US dollars in patent tax and research support fees were paid to Cornell University at one time. At that time, Bio-Rad Laboratories, Inc. signed an OEM and distributor agreement with DuPont. Subsequently, Bole Company introduced PDS- 1000/He gun in 1992. In China, the Institute of Biophysics of Chinese Academy of Sciences and Tsinghua University have also introduced new gun types in 1989 and199 respectively and applied for patents (China patents 89 109334 and 9 1207467). Different from the new hand-held gene gun, the desktop gene gun is too large and the experimental site is limited, so it can not be flexibly used in the field. High-pressure gas needs to be evacuated, and the compressor is noisy when working. Moreover, the high pressure pushes the particle bombardment, which makes the desktop gene gun limited to cell transplantation and cannot be used for living transplantation. The cost of each bombardment of the first generation gene gun is also very high, and the gold powder and rupture disk for controlling air pressure are very expensive.
The second generation gene gun appeared in 1996, and Bole Company introduced Helios handheld gene gun. This is the earliest handheld gene gun in history. The system drives gold particles prepackaged with DNA, RNA or other biological materials on the inner wall of a small plastic tube by adjustable helium pulse and injects them directly into cells. Compared with the first generation desktop gene gun, Helios handheld gene gun abandons the vacuum compressor and sacrifices part of the gas pressure, thus making it possible to transplant living animals, and can directly transplant the muscles and skin of living animals. Because of its small size, it is convenient for experimenters to carry around, which greatly broadens the application range of gene gun. In the following 10 years, Helios hand-held gene gun was widely used for gene transfer of monocotyledonous plants which were difficult to regenerate plants from protoplasts and insensitive to Agrobacterium infection. Before the gene gun, it was difficult for foreign DNA to pass through the organelles of the double-layer membrane after entering the cytoplasm. Gene gun technology is the most commonly used and effective DNA introduction technology in this field because of its high transformation frequency and good repeatability. Compared with the first generation desktop gene gun, the handheld gene gun can't penetrate the cell wall of mature leaves because of its low air pressure (only 100-600 psi), which affects the application range of transgenic plants to some extent, but it is complementary to the desktop gene gun, and Helios expands the application field of gene gun well.
Similarly, high-pressure gas is used to transfer genes, and the improvement of production technology makes the gene gun transfer from cells to living bodies, from desktop to handheld, and the application scope of gene gun is expanded step by step. The scientific research level of life science workers who first realized and actively tried to use gene gun in transgenic work has been greatly improved, which liberated the imagination of transgenic workers. The vigorous development of gene gun technology has been highly anticipated by the academic circles and regarded as the star of tomorrow in the field of genetic modification. However, people gradually found that the organs of living animals are much more fragile than skin and muscles. For example, the liver and spleen of living mice can only bear the pressure of 40psi at most, and under the impact of high-pressure gas of 100psi, the organs will be seriously damaged, leading to the failure of the experiment. However, too low air pressure can not make gene microcarriers have enough momentum to penetrate cells. The contradiction between gas pressure and particle transfer speed has become the bottleneck of the development of gene gun, which has been puzzling the R&D teams of major life science instrument manufacturers for the next 10 year.
Until 2009, Wealtec Company introduced GDS-80 low-pressure gene delivery system (also known as GDS-80 gene gun) (U.S. PatentNo. 6,436,709b1), which led the development direction of the third generation gene gun technology. GDS-80 hand-held gene gun skillfully starts from fluid dynamics and aviation dynamics, and uses helium or nitrogen to accelerate biomolecules to extremely high speed at low pressure to complete gene transfer, thus solving the contradiction between air pressure and particle speed from a new perspective. The ultra-low pressure (10-80 psi) of the third generation gene gun not only greatly increases the particle transport capacity without sacrifice, but also enables the gene gun to be successfully applied to animal living organ transplantation only at low pressure. Moreover, compared with the second-generation hand-held gene gun, the gene-carrying particles emitted by GDS-80 can penetrate the plant cell wall like the particles emitted by the desktop gene gun because of their large momentum, and complete the transformation. Prior to this, the first generation of desktop gene gun needed at least 1000-2000 psi of high pressure gas to complete this work. In the experiment of animal cells, especially living animals, it is not necessary to transfer biological particles with high momentum to the target body with the help of particle carriers (such as gold particles), which avoids the problem of foreign body residue in the target cell and greatly reduces the experimental cost. The low voltage conduction of the third generation gene gun greatly reduces the cell damage and the noise of gun bombardment, and effectively reduces the expensive cost of gold powder microcarriers in animal experiments. The preparation of GDS-80 gene gun "bullet" was also changed from dry method to wet method, which saved the drying time and simplified the process.