CN1036978C - Method for prepn. of tungsten bullet of gene gun for transfering external gene - Google Patents

Abstract

The present invention provides a preparation method of miniature biologic tungsten bullets for gene guns, which is characterized in that an ultrathin material bed is adopted to directly reduce yellow tungsten (WO3) by a low temperature dry hydrogen sublevel reduction method to prepare the miniature tungsten bullets for gene guns, wherein the thickness of the material bed is from1 mm to 10mm, the dew point of the hydrogen is from-70 DEG C to-20 DEG C, the flow rate of the hydrogen (l/min)/ yellow tungsten reduction amount (cm<3>) is from 0.05 to 1.0, the prereduction temperature is from500 DEG C to 650 DEG C with the time of 5 to 20 min, and the reduction temperature is [from]650 DEG C to 800 DEG C with the time of 5 to 20 min. The present invention has the advantages of low cost, uniform product particle size and polyhedral monocrystal particle pattern, wherein the average particle size is from1 to 2 mum, and the particles account for more than 90% in the range of average particle size (+/-)0.75 mum.

Description

The preparation method of tungsten bullet of gene gun for transfering external gene

The invention belongs to the category of genetic engineering and powder technology.

Gene gun technology is the transgenosis state-of-the-art technology that late nineteen eighties grows up in the genetic engineering, it is to utilize particle gun (or claiming particle gun, particulate launching device etc.) that the foreign gene that is coated on the miniature bullet (also claiming miniature year grain) is injected in plant, the animal or human's body intact cell with high speed, thereby reach the transmission foreign gene, the purpose of improvement plant, animal or reparation, regeneration tissue.Since gene gun technology than its also transgenic technology have its unique advantage in many aspects, thereby since 1987 are born, developed rapidly.Three patents about gene gun technology (EP0331 855A2 has successively declared in U.S. Cornell University from 1988,1988:US Patent Number 4,945,050 July 31,1990:US Patent Number 5,036,006 July 30,1991), E.I.Du Pont Company has released the PDS-1000 rifle in nineteen ninety, and Bole company released the PDS-1000/He rifle in 1992; Domestic Instite of Biophysics, Chinese Academy of Sciences and Tsing-Hua University have also released new variety of gun respectively at 1989 and 1991 and have applied for patent (Chinese patent 89109334 and 91207467).At present, particle gun commercialization at home and abroad.

Miniature bullet in the gene gun technology is actually that density is big, chemical stability good, and median size is the metal-powder of 0.4～2.0 μ m, as tungsten powder and bronze etc.The material of miniature bullet, particle size uniformity and shape etc. are the key factors that influences gene gun technology foreign gene transmission efficiency.Wide, in irregular shape little the launching of particle size distribution shows extremely inconsistent physical behavior (speed, direction, kinetic energy etc.) on the target plane, influence the effective performance of particle gun performance and the transmission efficiency of gene gun technology foreign gene.Though the rifle body of gene gun technology design has at present reached suitable level, the development of the stagnant rear lance of the research of miniature bullet is not seen as yet so far and is specialized in the report of particle gun with miniature bullet.Though E.I.Du Pont Company has product to sell, its technology is still maintained secrecy to external world.Used miniature tungsten bullet mainly is the special-purpose tungsten powder or the homemade general industry tungsten powder of E.I.Du Pont Company at present in the domestic gene gun technology, the former (1.8ml phial that costs an arm and a leg, 25 dollars of per 1 bottles), latter's particle size distribution is wide, out-of-shape, the particle that contains a large amount of non-three-dimensional shapes is difficult to satisfy the requirement of gene gun technology.

The objective of the invention is to the prepared sizes narrow distribution range, the particle gun of regular shape is with miniature tungsten bullet.

In theory, when the oxide compound of reduction tungsten was produced tungsten powder, growing up of tungsten particle can be thought what chemical vapors transport reaction (being called for short CVT) caused, and its reaction formula is:

Here WO_3-xBe a kind of protoxide, at high temperature volatile.Usually on thinner particle oxidation takes place, and on thicker particle protoxide by reduce deposition again.Thereby to obtain thin and the little tungsten powder of particle size distribution, and must control the carrying out of above-mentioned reaction process by control process parameters as far as possible, these processing parameters comprise reduction temperature, time, bed thickness, dew point of hydrogen etc.

Formation of the present invention is to adopt the ultra-thin bed of material, the low temperature dry hydrogen direct vat yellow tungsten of reduction method (WO stage by stage₃) produce the required miniature tungsten bullet of gene gun technology.Its processing parameter is (with reference to accompanying drawing): bed thickness 1mm～10mm, dew point of hydrogen-70 ℃～-20 ℃, hydrogen flowing quantity (1/min)/reductive yellow tungsten amount (cm³) 0.05～1.0, prereduction temperature (T_y) 500 ℃～650 ℃, reduction temperature (T_H) 650 ℃～800 ℃, prereduction time (t_y) 5min～20min, recovery time (t_H) 5min～20min.Reduction is carried out in horizontal chamber oven, can adopt to push away the boat mode continuously and also can adopt the discontinuous boat mode that pushes away.Discontinuous when pushing away the boat mode when taking, room temperature is to prereduction temperature heating-up time (t₁) 4min～10min, the prereduction temperature is to reduction temperature heating-up time (t₂) 1min～2min.Adopting the inventive method and the prepared miniature tungsten bullet of processing parameter is the polyhedron single crystal particle, and mean particle size is 1～2 μ m.Because the present invention adopts the ultra-thin bed of material, low temperature dry hydrogen short period of time method of reducing, guarantee all chargings reduction simultaneously,---volatilization---restores deposition process to have suppressed the oxidation in the reduction process effectively, and prepared miniature tungsten bullet can improve significantly than general industry tungsten powder particle-size homogeneity.

The invention has the advantages that preparation cost is low, product granularity is even, and granule-morphology is the polyhedron monocrystalline, and mean particle size is 1～2 μ m, and mean particle size ± 0.75 mu m range endoparticle accounts for more than 90%.

The present invention is further described below in conjunction with accompanying drawing.

Fig. 1 is the synoptic diagram of technology of the present invention, and abscissa t is the time, and unit be minute that ordinate T is a centigradetemperature, puts T_HBe reduction temperature, T_yBe prereduction temperature, corresponding t_HWith t_yBe soaking time.

Embodiment:

Adopt the inventive method and two groups of processing parameters to prepare particle gun with miniature tungsten bullet in the discontinuous boat mode that pushes away, processing parameter such as table 1, used yellow tungsten particle degree is 0.51 μ m.The comparison such as the table 2 of miniature tungsten bullet of miniature tungsten bullet of the present invention and E.I.Du Pont Company and general industry tungsten powder.Particle size uniformity characterizes with the particle percentage ratio in mean particle size ± 0.75 mu m range in the table.Table 1 implementing process parameter

	Processing parameter A	Processing parameter B
			Bed thickness (mm)	5	2
Dew point of hydrogen (℃)	-31	-31
			Hydrogen flowing quantity (1/min)/yellow tungsten amount (cm3)	0.4	0.6
The prereduction temperature Ty(℃)	650	600
			Reduction temperature TH(℃)	750	750
Prereduction time ty(min)	15	10
			Recovery time tH(min)	15	20
Heating-up time t1(min)	10	10
			Heating-up time t2(min)	2	2

The comparison of the miniature tungsten bullet of table 2 embodiment, the miniature tungsten bullet of Du Pont and common homemade industrial tungsten powder

	Parameter A	B parameter	Du Pont's powder	The industry tungsten powder
					Granularity (μ m)	1.63	1.54	1.67	1.9
Particle size uniformity	90％	＞90％	90％	60％
					Granule-morphology	The polyhedron monocrystalline	The polyhedron monocrystalline	Unsetting	Unsetting

Claims (1)

1, a kind of particle gun preparation method of miniature organism tungsten bullet, it is characterized in that adopting the ultra-thin bed of material, the low temperature dry hydrogen direct vat yellow tungsten of reduction method stage by stage, produce the required miniature tungsten bullet of particle gun, bed thickness 1mm～10mm wherein, dew point of hydrogen-70 ℃～-20 ℃, hydrogen flowing quantity (1/min)/reductive yellow tungsten amount (cm³) be 0.05～1.0,500 ℃～650 ℃ of prereduction temperature, time 5-20min, reduction temperature are 650 ℃～800 ℃, the recovery time is 5～20min.

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