TW201139674A - Low pressure accelerated gene delivery device and barrel structure thereof - Google Patents

Abstract

A barrel structure of a low pressure accelerated gene delivery device including a pressurized chamber and a spray nozzle is described. The spray nozzle is connected to the pressurized chamber, wherein the inside diameters of the spray nozzle front-terminal and the pressurized chamber are the same, and the outside diameters of the spray nozzle and the pressurized chamber are the same. The spray nozzle includes an interior contour having a converging part, a diverging part and a throat part between the converging part and the diverging part. The converging part is extending from the spray nozzle front-terminal to the throat part and the diverging part is extending from the throat part to the spray nozzle back-terminal. The connection between the converging part and the pressurized chamber is curve-shaped. The diverging part linear-divergently extends to the back-terminal of the spray nozzle. The diverging part can also parabolic-divergently extend to the back-terminal of the spray nozzle.

Description

Vi.doc/n 201139674 VI. Description of the Invention: [Technical Field] The present invention relates to a low-pressure airflow accelerated gene delivery device and a tube-preventing structure thereof, and in particular to a simple structure and easy to cast and can be A device that reduces gas turbulence, has high gene transfer efficiency, and a tube-preventing structure. [Prior Art] With the discovery of genetic material and the rapid development of genetics, humans have been able to use genetic genetic techniques to use foreign genes to alter the expression of a cell, or an individual. With the advancement of biotechnology, the genetic material deoxyribonudeie acid (DNA) of other organisms has been transferred into another organism to change the traits of the organism, which has been widely used in basic research and agricultural crops. Improvements, such as increasing the cold resistance, insect resistance and changing nutrients of crops, etc. In recent years, technology for gene transfer has also begun to be used in the medical treatment of human and animal diseases, including gene therapy and genetic vaccines. The successful application of gene transfer in the field of medical prevention and treatment will bring about major medical breakthroughs for many genetically related diseases. In recent years, the newly developed gene for the use of physical gene transfer technology has taken a big step forward in the application of clinicalization of gene transfer technology. The method utilizes gold particles carrying biological substances (such as DNA) to cause biological substances to enter biological cells by means of firing, thereby achieving the purpose of gene transfer. It has been used in many research and development fields, such as plant systems,

S 4 Ting, doc/n 201139674 Mammalian somatic cells, gene therapy, and even the recent DNA vaccine research system. At present, there is a novel gene grab designed by the theory of gas dynamics, which can use a liquid (such as nitrogen, helium or air) to contain a liquid solution containing biological substances (such as DNA or RNA or protein) under low pressure. Instantly accelerates to very high speed (greater than 200 m/s), so it is not necessary to use carrying particles (such as gold, tungsten particles, etc.) to allow biological substances to pass through the surface structure or cell membrane of the organism, into the cytoplasm or nucleus. Biological cells express special foreign proteins, or genes are transferred to produce new biological functions. The current application areas include animals, plants and cell lines. However, the complicated structure of the gene grab used at present is highly difficult to cast, and gas turbulence is easily generated in the accumulator chamber of the gun barrel structure, which greatly reduces the gene transfer efficiency. SUMMARY OF THE INVENTION In view of the above, an object of the present invention is to provide a low-pressure airflow accelerated gene delivery device and a tube-preventing structure thereof, which have the characteristics of easy casting, and the tube-preventing structure can effectively eliminate the generation in the pressure storage tank. Gas turbulence phenomenon. The present invention provides a pipetting structure for a low pressure gas flow accelerated gene delivery device comprising a pressure storage compartment and a nozzle. The nozzle includes a front end of the nozzle and a rear end of the nozzle, and the front end of the nozzle is connected to the accumulator chamber, wherein the inner diameter of the front end of the nozzle is the same as the internal control of the accumulator chamber, and the outer diameter of the nozzle is the same as the outer diameter of the accumulator. The nozzle has an inner contour, the inner contour including a tapered portion, a gradual extension portion, and a throat between the tapered portion and the gradual portion, wherein the tapered portion extends from the front end of the nozzle...doc/n 201139674^2!:: throat Extend to the back end of the nozzle. In particular, the curvature of the semi-circular arc of the gradual end is half (four), the radius of the second front R, and the radius of the throat is the other half of the radius of the iliac crest is half-column from the throat to the ancient B system 々<;^<2々. Moreover, the progressive diameter is a gradual function, and (2) the expansion extends to the rear end of the mouth, where the radius of the progressive portion DD=rT+(rE.rT)x/L〇Γε h, The gradual extension is the radius of the back end of the nozzle. The length and the Γτ are the radius of the throat. Improvement = the throat is parallel and the short straight section is designed. 1 Mandrel ^ The mouth to the throat t is the self-recording with the linear taper function For: & 'The diameter of the tapered part is a decreasing function, its linearity

Dc=^i^j~ rT)x/Lc Position: The position of the tapered portion from the pressure chamber of the animal, which is the + diameter of the tapered portion, and the radius of the nozzle tip. The length, rT is the throat radius, A. The present invention further provides a 201139674idoc/n tube structure for a low pressure airflow accelerated gene delivery device, which includes a pressure storage chamber and a nozzle. The nozzle includes a front end of the nozzle and a rear end of the nozzle, and the front end of the nozzle is connected to the accumulator chamber, wherein the inner diameter of the front end of the nozzle is the same as the inner diameter of the accumulator chamber, and the outer diameter of the nozzle is the same as the outer diameter of the accumulator chamber. The nozzle has an inner contour, the inner contour including a tapered portion, a progressive portion, and a throat between the tapered portion and the gradually extending portion, wherein the tapered portion extends from the front end of the nozzle to the throat, and the progressive portion extends from the throat to The back end of the nozzle. In particular, the tapered portion and the pressure accumulating chamber end are joined by a circular arc, and the relationship between the radius of curvature of the circular arc & and the radius 〇 of the tip end of the nozzle is 艮22〇. Further, the relationship between the curvature radius Rt of the throat and the radius of the throat is < Rt < 2rr. Moreover, the gradual extension is extended from the throat with a bell-shaped curve to the rear end of the nozzle, wherein the exponential function of the bell curve of the gradual extension is as follows: r = rT + b (l-e'ax ) a = -1/ Ld* In [\-{rE-rT)lb] b>rE-rr where x is the position of the gradual portion from the throat, r is the radius at the position X of the gradual portion, and Zz) is the gradual portion The length is the radius of the throat, r is the radius of the back end of the nozzle, 6 is the parameter defining the shape of the gradual part, and α is the curvature calculated according to the value. In one embodiment, the throat is designed as a parallel short straight section to improve the atomization of the sample liquid. In an embodiment, the taper is a point where the straightening of the pressure tank is linearly extended to the position of the throat, and the diameter of the taper is a decreasing function, and the linear function is:

Dc^Yi-(ri~ rT)x/Lc 7 201139674 /i.doc/n is the length of the front end of the nozzle, the length of the throat, the radius of the throat, this month is another proposed - low-pressure airflow accelerated gene delivery device , 1 W , # 9 The buffing device C includes a control valve and the starting device/set 1 = 2 is disposed between the starting device and the accumulator. Gas is connected. The barrel housing is provided with a phased nozzle to allow the squirting to steadily engage the firing system. In the case of the above-mentioned apparatus, the above-mentioned apparatus further includes an annular spacer ‘sleeves&' where the annular spacer has at least a gap. Heart:::intermediate =:=, feed hole position: is a particulate biological material or; 'and === material feed t embodiment, the above device further includes a feeding device, and is set at t The towel of the embodiment described above is an electromagnetic-mechanical valve. Grab the two 进 injection: ΐ has the same outer diameter as the outer diameter of the inspection, so: the construction is easy to enter the casting. In addition, the inner end of the nozzle is connected to the accumulator chamber, and the inner rib is the same as the inner diameter of the accumulator chamber. The retracting portion and the f-ball chamber end are connected by a round fox, so that the gas can be effectively eliminated from the pressure (4). Correction of the parallel short straight section of the throat, in addition to the placement of the sample, can enhance the atomization effect. In order to achieve an even exit velocity, the inner wheel of the progressive section

S ^i.d〇c/n 201139674 Chad is designed as a bell. On the other hand, the annular spacer is placed at the rear end of the nozzle to maintain a fixed distance between the low-pressure gas gripping speed gene delivery device and the target to be fired' to reduce the probability of shooting due to gene grabbing. The distance between the targets is offset by an error, and the annular spacer has at least one gap to prevent the strong airflow from injuring the target to be fired. The above and other objects, features, and advantages of the present invention will become more apparent and understood. [Embodiment] FIG. 1 is a schematic diagram of a low-pressure airflow accelerated gene transfer device system according to an embodiment of the present invention. Fig. 2A is a schematic view showing the structure of a barrel of an embodiment of the present invention. g] 3 is shown as a partial enlarged view of the circle in Figure 2A. Hereinafter, referring to Fig. 2, Fig. 2A and Fig. 3, the low pressure gas flow accelerated gene delivery device system 1 mainly includes a gas supply device 102's pipetting structure 104 and a firing skirt 1〇6. The gas supply unit 102 includes a gas cylinder 112, a pressure regulator I" and a line 116, wherein the line 116 is connected between the pressure regulator 114 and the firing device 106. The gas used is supplied by a gas cylinder 112, and the gas in the gas 112 may be nitrogen, helium or other gases. The quality of hernia is relatively small, and it is not ideal for injuring cells. It is a very desirable gas, but it is more expensive. Some plants and plants with thicker cuticles are still not used. However, when using a gene gun system in some biological systems that are easier to pass, it is sufficient to use nitrogen w-doc/n 201139674 gas. , 1 to set the required pressure ^ 先 first through a pressure regulator Μ S structure 1 〇 4 includes the pressure accumulating face 122 and the nozzle 124. The pressure storage tank 122 steel or its three materials: ϊ! metal or plastic steel, for example, the alloy, the easy-to-convert excellent gun: with the alternative nozzle, the diameter and the diameter of the nozzle, the nozzle 124 is wrapped in a , the nozzle rear end i24b, and the nozzle front end i24a are connected to ^. The inner diameter of the end portion 124a of the nozzle is opposite to the inner diameter of the accumulator chamber 122, which can eliminate the disturbance generated between the gas flow storage tank 122 and the nozzle, which is said to be 'because the nozzle front end 124 & , I I and the accumulator 122 and the gas supply device ι〇2 line outlet 11 ^, there is a phase _ 彳 彳 ' and thus the junction between the two will not have a dead angle or corner f β ' so when the gas is supplied from the gas When the device 102 enters the accumulator compartment 122 and enters the nozzle tip end ma by the accumulator 脍m, there is a gas turbulence present. The nozzle I24 has the same outer diameter as the pressure accumulating chamber m, so that the pipe grabbing structure 104 is fresh and easy to cast. The material used for the nozzle m is a biocompatible metal such as an aluminum alloy, stainless steel or alloy thereof. The nozzle 124 has an internal contour that includes a tapered portion 126, a tapered portion 128, and a throat 130 between the tapered portion ι26 and the tapered portion ι28. The tapered portion 126 extends from the nozzle front end 124a to the throat portion 13b, and the progressive portion 128 extends from the throat portion 130 to the nozzle rear end 124b. In addition, the pipe grabbing structure 104 may further include a pipe grabbing casing 134 which is engaged with the firing device 1〇6 by threading the pipe casing 134 to fix the pipe grabbing structure 104, and may make the gene gun more beautiful. 201139674 The firing device 106 includes a control valve 118 and an activation device 136, wherein the control valve 118 is disposed between the activation device 136 and the accumulator compartment 122. After passing through the pressure regulator 114 and passing through the line 116, the gas is first controlled by the starting device 136 to enter the accumulator compartment 122 of the barrel structure 1〇4 via the control valve 118. The control valve 118 is, for example, a solenoid valve or a mechanical valve. In one embodiment, control valve 118 is controlled via a controller (not shown). When using the low pressure gas flow accelerated gene delivery device of the present invention, the sample is first introduced from the feed port 132, for example, by feeding the sample using any suitable feeding device (not shown), for example Feeding is carried out using a needle, pipette or automatic delivery system. The feeding device in the structure of the present invention is not limited to a single-technology or device, and any currently known structural device or method can be applied. The ya% spacer 131 is placed at the outlet of the nozzle rear end 124b, so that the low-speed, air-accelerated gene delivery device and the target to be fired are secured, and the solid-pitch spacing is called low-low-pressure airflow accelerated gene delivery. The error caused by the offset of the low-pressure airflow-accelerated base delivery device and the target to be fired when the device is in two shots. The annular spacer i3i has a defect 140, and gas can flow out of the notch 140 to prevent the strong milk & damage to the target to be fired. During operation, 'set the pressure to 1 1Μ to the appropriate one. For this purpose, the low-pressure airflow accelerated gene is cast and the target of the target is shot Μ + / contains biological substances (such as RNA, DNA or protein) 'cold liquid Put in the feeding device. At the time of shooting, the gene grabs the structure. 11 201139674.1 - The device 106 opens the control valve 118 via a starting device 136 to allow gas to flow into the accumulator chamber 122, generating a high velocity gas stream to drive the sample solution, and successfully hitting the target through the nozzle 124. In particular, the nozzle 124 is designed as a tapered tapered nozzle, i.e., it includes at least a tapered portion 126, a tapered portion 128, and a throat portion 13 between the tapered portion 126 and the tapered portion 128. The purpose of the throat 13 is to create a continuous curve between the taper = 126 and the progressive portion 128 to allow the gas to flow smoothly. In addition, in order to enable the solution particles to operate at a higher speed, the progressive portion 128 designed by the present invention allows the gas to reach supersonic speed, and the solution particles are closer to the speed of sound, so that the metal can be carried without the need for particles to penetrate. The throat 130 of the nozzle 124 is designed with a curved inlet, as shown in FIG. 3, which allows the gas to be distributed more uniformly at its 丨σ, so that the solution particles are evenly distributed, and the towel is not distributed in certain areas, resulting in The phenomenon of cell death. Moreover, the pressure of the gas at the outlet can be brought close to one atmosphere, avoiding damage to the target cell tissue. In particular, the emergency structure of the low-pressure airflow-accelerated gene delivery device of the present invention has the following special structural design: (-) Designing the throat of the tapered portion, the progressive portion, and the throat 130: rT<Rt<2rT The section 128: 〇<1<15 degrees to 2 sheets of 邛I28 is designed as a conical tube having an angle of expansion. After the airflow passes through the 126 and the throat 130, it gradually passes through the gradual expansion. Shoot out. In Fig. 3, Rt represents the radius of curvature of the throat 130, and rT is the throat. The radius of P 13〇, * is the progressive part 128 and the central axis (virtual

The angle of 12 S *_doc/n 201139674 line). In order to change the cover σ, add a parallel short straight tube to get in the throat! 3〇 The effect of atomization. Sample. ° Liquid hanging drops, also have improved both the shrinkage and the face test are connected by a circular arc. The arc curvature is connected, and this can make there is no dead angle between the two: the joint. Dan a corner thus eliminates gas turbulence (3) contour design of the gradual extension _ towel 'the inner wheel of the ship's 128 is straighter =: gradual / = 128 is extended from the throat 130 by a straight line extension # 2A)) and the diameter of the progressive portion 128 is an increasing function:

^ D = rT + (rE - rT) x / LD where x is the position of the progressive portion 128 from the throat 130, Dd is the radius at the position x of the progressive portion 128, and Ld is the length of the progressive portion 128 = rT is the radius of the throat 13 Γ, and Γ ε is the radius of the outlet of the nozzle 124. In another embodiment, the inner contour of the progressive portion 128 is a bell-shaped gradation, as shown in Figure 2A. That is, the progressive portion 128 extends from the throat 13〇 in a bell-shaped curve to the rear end of the nozzle 124. The exponential function of the bell curve of the progressive portion 128 is as follows: r=rT+b (\-eax) 13 vi.doc/n 201139674 a=-l/LD*\n[\.(rE_rTyt>] fτ where x is the position of the progressive portion 128 from the throat 130, and r is the position X of the progressive portion 128 The radius '4' is the length of the progressive portion 128, the radius 130 of the throat, the radius of the rear end of the nozzle 124, & is the parameter defining the appearance of the progressive portion 128, "the curvature calculated according to the value of 6" The larger the value of 'b, the smaller the slope of the curve is, which is closer to the straight line; the smaller the value of b, the larger the slope of the curve is, which is closer to the bell shape. In addition, the inner contour of the tapered portion is linearly tapered. In a preferred embodiment, the tapered portion 126 of FIGS. 2A and 2B is a point where the diameter of the tapered portion 126 is a decreasing function from the position where the pressure accumulating chamber 122 is tapered in a straight line to the position of the throat portion 130. The linear function is:

Dc = n - (ri - rT) x / Lc where x is the position of the tapered portion 126 from the load storage chamber 122, Dc is the radius at the position X of the tapered portion 126, and Lc is the length of the tapered portion 126 rT is the radius of the throat 130 and η is the radius of the front end of the nozzle 124. Further, the tapered portion 126 and the progressive portion 128 of the nozzle 124 can also be designed according to the following description: Assuming that the flow field is an isentropic flow, the ratio of the outlet area VIII to the throat area At is:

Ae At

/-1 ~2~ Me2 r+i 2〇M) where Me is the Mach number of the exit (Mach number = gas flow rate / speed of sound). If the required Mach number and Ae are selected, the throat area At and the 201139674 are obtained. The full pressure p〇 of the nozzle inlet is required.

Po = P 1-l·

For the flow field from the gas cylinder to the pressure storage tank 122, the pressure of the pressure accumulation 122 is its outlet pressure, so the pressure of the cylinder 112 is required.

Since a certain speed is required to enable the blowing of the sample solution, the Mic speed (Min) entering the pressure storage tank 122 is selected to obtain the desired cylinder pressure. In addition, in order to maintain a constant pressure in the accumulator chamber 122, the mass discharged from the nozzle 124 must be equal to the mass of the inflow. Min={pAV).m=(pAY)〇w

From the above formula, the population area required for the flow storage tank 122 can be obtained. Experiments The following experiments were conducted using actual experiments. In this experiment, the sample for the position of the feed hole and the delivery efficiency was 16/i 1

Coomassie Blue, Fig. 4A shows the picture of Fig. 4B; Photograph of the input, 201139674, photo of the idoc/n voyage input, Fig. 4D shows the feed hole from the sample = part The photograph of the input _, green in Fig. 4E, is a photograph of the sample taken from the feed hole 7.5 cm from the throat. Referring to Figures 4A-4E, the results of Figures 4A-4E show that the Wei-injection is close to the throat, and the penetration is also better. YANG Sample Solution Preparation The preparation procedure of the sample solution, depending on the biological substance contained, has different preparation details, and thus is not limited to a single technique or procedure. Any known art or method can be applied. In general, depending on the nature of the biological substance contained, the biological substance is dissolved in a suitable solution without adding metal-carrying particles, and is obtained by appropriate treatment. Preparation of DNA solution The DNA plastid (Plasmid) pEGFP-N2, which constructs the complete fluorescent protein expression system, is dissolved in sterile TE buffer and can be directly placed in the feed device. 'For low-pressure airflow accelerated gene delivery device The bombardment. Preparation of TE aqueous solution containing PEI

PEI (p〇lyethyleneimine, Sigma P3143, 50% (w/v) in water) was dissolved in sterile distilled water to make an i2.5% (w/v) aqueous solution, and then diluted with TE buffer to 〇.〇〇〇〇 i% (〇.ing/pl), that is, a TE aqueous solution containing PEI is prepared.

S 16 201139674, oc/n Preparation DNA-PEI complex The preparation of the complex is completed by uniformly mixing the DNA with the PEI aqueous solution at a ratio of PEI:DNA=lng:lpg. Preparation of DNA-spermidine-CaC12 complex solution Aspirate 5 ul (lug/ul) of plastid DNA (pBI121, pGR, pCG) and 20 ul of 0.1 M spermidine in an ultrasonic wave to homogenize. Add 50 ul of 2.5MCaC12 shake mix. After standing for 5 minutes, 140 ul of 70% alcohol was added and centrifuged at 12,000 rpm for 2 seconds. After carefully removing the supernatant, add 14 μl of absolute alcohol (100%) without damaging the DNA pellet, and centrifuge at 12,000 rpm for 2 seconds. After removing the supernatant, the DNA pellet was reconstituted with 50 ul of absolute alcohol. Preparation of EGFP protein solution The purified fluorescent protein EGFP was dissolved in sterile water and directly placed in a feeding device for bombardment of a low-pressure airflow accelerated gene delivery device. Experimental design of bombarding mouse skin with low-pressure airflow accelerated gene delivery device. Select 5-week C57BL6 mice, remove the abdominal hair of the mouse with a razor, and expose the smooth aqueous tissue tissue, DNA aqueous solution, DNA-PEI complex aqueous solution or EGFP protein solution. After the low-pressure airflow accelerated gene delivery device was placed, the exposed skin of the mouse was bombarded with a low-pressure airflow accelerated gene delivery device. The skin of the abdominal skin of each mouse was about four bombardments, and the volume of each solution was 20 μl. The mice were sacrificed the next day after the gene bombardment, 17 vi.doc/n 201139674 to remove the skin tissue, and the removed skin tissue was treated directly under a fluorescent microscope. The low-pressure airflow accelerated gene delivery device bombarded the mouse skin wide test results directly bombarded with DNA liquid solution with 20, 30, 40 and 50 psi pressure helium, with a 10 mm inner diameter of the tube to carry out low-pressure airflow acceleration gene The bombardment of the delivery device was 20 μl per solution, containing 3 pg of DNA, and the fluorescent appearance of the bombarded abdomen skin of the mouse was observed under a fluorescence microscope. Figures 5A-5D show the results of a 40x magnification of a fluorescent microscope, in turn, at 2, 30, 40, and 50 psi. From the experimental results, it can be seen that the skin tissue bombarded at 3 psi and 40 psi has the best green fluorescence performance. The aqueous solution of PEI-DNA complex was bombarded with helium gas at a pressure of 30 psi, and bombarded with a low-pressure airflow accelerated gene delivery device with a barrel having an opening diameter of 10 mm. Each solution was a 2 μμ1 aqueous solution of PEI-DNA complex. 1|Xg DNA was contained, and the fluorescent expression of the bombarded abdomen skin of the mouse was observed under a fluorescence microscope. As shown in Fig. 6, when PEI was added to the solution, the DNA dose was lowered to lpg, and the fluorescence was as fluorescent as that of the aqueous solution containing 3 pg of DNA per hair. Bombardment with EGFP protein 201139674 d ; /r.doc/n Low-pressure airflow accelerated gene delivery device with a helium 5 psi pressure shot containing 5/zg EGFP protein per shot. Fluorescence microscopy

Bombardment of the skin of the mouse's abdomen. As shown in Fig. 7-8 to Fig. 7C, the radiance microscope was observed at 40 times magnification to observe the abdominal skin of the mouse, and it was observed that the EGFP protein was successfully and uniformly injected into the epidermal layer. /T bombards the petal 〇f Oncidium Ramsey with a low-pressure airflow-accelerated gene delivery device. The helium gas with a pressure of 50 psi is used to bombard the low-pressure airflow-accelerated gene delivery device with a tube with an opening diameter of 4.5 mm. The solution was a DNA-spermidine-CaC12 complex aqueous solution containing igg DNA, which was 3 cm away from the target, and stained with guS tissue on the next day. The coloration of GUS was observed after 48 hours of reaction in the dark room. . It can be seen from the foregoing experimental results that the low-pressure airflow accelerating gene delivery device used in the present invention does not cause serious damage to the target cell tissue. Moreover, the low-pressure airflow accelerated gene delivery device of the present invention can directly transfer genetic DNA or other biological substances into the interior of the cell, thereby avoiding the consumption of barriers hindered by many tissues or cells, and successfully achieving gene transfer or biological substances. The purpose of delivery. In general, the present invention has at least the following advantages: 1. In the low-pressure airflow accelerated gene delivery device proposed by the present invention, the nozzle of the pipe-collecting structure has the same outer diameter as that of the pressure storage tank, making the pipe-sucking structure easy Casting. 19 201139674. 2, the low-pressure airflow-accelerated gene delivery device proposed by the present invention has a same inner diameter at the junction of the sigma, the sigma device and the animal pressure chamber, and the tapered portion and the accumulator chamber are in a circular arc phase. The connection can eliminate the phenomenon of spoilage when the gas enters the accumulator. 3. The inner contour of the gradually extending portion of the pipe grabbing structure of the present invention is a linear expansion or a bell expansion, and such a design can increase the acceleration of the emitted gas and make the emitted gas have a high uniformity. 4. The low pressure airflow accelerated gene delivery device of the present invention has an annular spacer which maintains a fixed spacing between the low pressure airflow accelerated gene delivery device and the target to be shot to reduce the shooting error. In addition, since the annular spacer of the low-pressure airflow-accelerated gene delivery device proposed by the present invention has at least one notch, it is possible to prevent the strong airflow from injuring the target to be fired. Although the present invention has been described in the above preferred embodiments, it is not intended to limit the present invention, and those skilled in the art can make some modifications and refinements without departing from the spirit and scope of the present invention. The scope of the invention is defined by the scope of the appended claims. [Brief Description of the Drawings] Fig. 1 is a schematic view showing a low-pressure airflow acceleration type gene delivery device according to an embodiment of the present invention. earth

2A and 2B are diagrams showing the structure of the pipe grabbing structure of the embodiment of the present invention. FIG. 3 is a partial enlarged view of the circled portion of FIG. 201139674 doc/π Pie Chart Figure 4A is a photograph of the sample taken from the feed hole above the throat. Figure 4B is a photograph taken from the sample hole from the throat of 1.2 cm from the throat. Figure 4C is a photograph of a sample taken from a feed hole 2.5 cm from the throat. Figure 4D is not a photograph of a sample taken from a feed hole 5 cm from the throat. The inset is shown from a sample of 7.5 cm from the throat of the throat into a 20 psi helium bomb with a helium bomb of 30 psi with a helium bombardment of 4 〇 pSi with a helium bomb of 50 psi. Magnification 4 times under the microscope, observation results obtained by hitting the abdominal skin of the mouse Figure 5B is a magnification of 4 times under a fluorescent microscope, and the observation result obtained by hitting the abdominal skin of the mouse is shown in Fig. 5C, which is magnified 4 times under a fluorescent microscope, and the mouse abdomen is hit. Observation Results of Skin Figure 5D is a magnification of 4 times in a fluorescent microscope. The observation results obtained by hitting the abdominal skin of a mouse are shown in Fig. 6. The magnification is 4 times in a fluorescent microscope under the microscope, and the abdomen is bombarded with a helium gas of 30 psi. Figure 7A shows the results of the observation with EGFP τ*, and the area is observed under a fluorescent microscope (40x magnification). Figure 7B shows the bombardment with EGFP protein. Under the fluorescence microscope (magnification 40 times) Guanjiamu β house milk skin tissue', Jinshen stretch results (delivery area Γ 21 201139674 'doc/n Figure 7C shows the bombardment of mouse skin tissue with EGFP protein, in fluorescent The results were observed under a microscope (magnification 40 times) (delivery area 2). Figure 8A shows the observation of staining with GUS tissue after bombardment of the petals of Oncidium with alcohol (no DNA), 50 psi of helium. Fig. 8B is a view showing the observation of GUS tissue staining with the pBIU1 solution and 5 psi of helium gas after bombardment of the petals of the heart. Figure 8C shows the bombardment of the helium gas with pRG solution and 5 psi. The observation results of GUS tissue staining after heart petals. ~ Lantu 8D is the observation result of GUS tissue staining with PCG solution '50 psi of helium gas bombarded with GUS tissue. ·Lan [Main component symbol description] 100: low pressure airflow accelerated gene Delivery apparatus 102: gas supply device 104: grab the tubular structure 106: the firing device 1〇6 112: cylinder 114: pressure regulator 116: line 118: valve 122: pressure chamber 124: Zuizui 124a: tip of the nozzle

S 22 201139674 doc/n 124b : nozzle rear end 126 : taper 128 : taper 130 : throat 132 : feed hole 134 : barrel housing 136 : starting device

Ae: cross-sectional area of the end of the progressive portion 128

At : throat 130 cross-sectional area

Rt: radius of curvature of the throat 130 rT: radius of the throat 130 Θ : angle between the progressive portion 128 and the central axis 23

Claims (1)

201139674 doc/n VII. Patent application scope: 1. A barrel structure of a low-pressure airflow accelerated gene delivery device, comprising: a pressure storage tank; and a nozzle comprising a nozzle front end and a nozzle rear end, the nozzle front end Connecting to the accumulator chamber, wherein an inner diameter of the front end of the nozzle is the same as an inner diameter of the accumulator chamber, the nozzle has an inner contour, wherein the inner contour comprises: a tapered portion; a gradual portion; and a throat a portion extending between the tapered portion and the tapered portion, wherein the tapered portion extends from the front end of the nozzle to the throat, the progressive portion extending from the throat to the rear end of the nozzle, wherein the tapered portion The portion is connected to the end of the pressure accumulating chamber by a circular arc, and the relationship between the radius of curvature R! of the circular arc and the radius r/ of the front end of the nozzle is 2rz, and the radius of curvature Rt of the throat and the radius of the throat The relationship between the plants is 7·< Rt < 2 Plant 2 &quot; ' The gradual extension is from the throat extending in a straight line to the rear end of the nozzle, and the diameter of the gradual extension is an increasing function, as follows: DD =rT+(rE- rT)x/L〇 where x is calculated from the throat The position of the gradation portion, Dd is the radius at the position X of the gradual portion, LD is the length of the gradation, rT is the radius of the throat, and rE is the radius of the rear end of the nozzle. S 24 201139674.doc/n 2. The barrel structure of the low-pressure airflow accelerated gene delivery device as described in claim 1 wherein the throat is designed as a parallel short straight section to improve the atomization effect of the sample liquid. . 3. The pipetting structure of the low-pressure airflow-accelerated gene delivery device according to claim 1, wherein an angle Θ between the gradual portion and the central axis of the nozzle in the inner contour is greater than a twist and less than 15 degrees. 4. The pipetting structure of the low-pressure airflow-accelerated gene-reporting device as described in claim 1 wherein the tapered portion is tapered from the pressure storage to the throat, the tapered portion The diameter is a decreasing function, and its linear function is · Z) c - τf) x / Lc where X is the position of the tapered portion from the pressure accumulating portion 'Dc is the radius at the position X of the tapered portion, Lc is the length of the tapered portion, & is the radius of the throat, and ri is the radius of the tip end of the nozzle. 5) a pipetting structure of a low-pressure airflow accelerated gene delivery device, comprising: a pressure storage tank; and a nozzle including a nozzle front end and a nozzle rear end, the nozzle front end being connected to the pressure storage tank, wherein the nozzle The inner diameter of the front end of the nozzle is the same as the inner diameter of the pressure accumulating chamber, and the outer diameter of the nozzle is the same as the outer diameter of the pressure accumulating chamber. The nozzle has an inner contour, wherein the inner wheel includes: a tapered portion; a gradual extension; and 25 201139674 d〇c/n between the tapered portion (four) and the gradual extension portion, which is extended to the nozzle ^ (four) portion, the gradual extension portion from the throat portion of the arc: The constricted portion is connected with the accumulating face end by a round fox, and the circle says that the radius of the front end of the nozzle and the __ of the nozzle is RI is "d, the radius and the radius of the throat" The relationship between the mouth and the mouth: Gradual:: The exponential function of the bell curve extending from the throat with a bell-shaped curve extending to the end of the nozzle is as follows: r=rT+b (\-eax) a~ -1/Ld* In b'^&gt;y£-rj· where ^ is the position of the gradual portion from the throat, r is the gradual portion: half at the vertical X , &amp; is the length of the gradual extension, &amp; is the radius of the throat half ' k '~ is the radius of the nozzle back end, 6 is the parameter defining the shape of the gradual extension, and α is calculated according to the value of 6 6. The pipetting structure of the low-pressure airflow accelerated gene delivery device according to claim 5, wherein the throat is designed as a parallel short straight pipe section to improve the atomization effect of the sample liquid. The pipetting structure of the low-pressure airflow-accelerated gene delivery device described in claim 5, wherein the tapering portion is a line that is tapered from the end of the pressure-storing chamber to the position of the throat, the tapering The diameter of the part is a decreasing function 'the linear function is: S 26 201139674 _ doc / n Dc = n - (ri - rT) x / Lc where x is the position of the taper from the tank, Dc哕 is the radius of the taper position X, Lc is the length of the taper, the radius of the throat, ij is the radius of the front end of the nozzle. τ...μ 8. a low-pressure airflow accelerated gene delivery device, Including a barrel structure as described in one of claims 7 to 7; a firing device comprising - a control port and a start I, the control valve being disposed between the activation device and the pressure storage tank; ^ a gas supply device coupled to the firing device; and - a barrel housing 'The pipe casing is a mosquito mosquito, and the nozzle mouth is firmly connected with the firing device. 9. The low-pressure airflow accelerated gene delivery device according to claim 8 of the patent application, further includes a feed hole, setting Between the gradual extension of the throat of the eclipse, the feed hole can be delivered with a biological material (10), a metal particulate biological material or a biological material mixed with liquid and metal particles. 10. The flow reduction accelerating gene delivery device according to item 9 of the scope, further comprising a feeding device installed at the feed hole. 11. As described in item 8! The low-pressure airflow accelerated gene delivery device's control side is an electric or mechanical valve.