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Nucleic acid synthesis solid phase carrier, preparation method, nucleic acid synthesis device containing the carrier, and nucleic acid synthesis method

A solid-phase carrier and nucleic acid technology, applied in biochemical equipment and methods, DNA preparation, biochemical cleaning equipment, etc., can solve the problems of inconvenient synthesis operation, reduce the amount of reagents, facilitate the synthesis method, and avoid plastic pollution Effect

Active Publication Date: 2021-06-04
艾维迪亚(深圳)医疗科技有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0003] The inventors have found that when the existing plastic synthesis column is used to complete nucleic acid synthesis, the specifications of the plastic synthesis column are fixed, and only a fixed scale of nucleic acid fragments can be synthesized on the same equipment, such as 50nmol, 100nmol, 200nmol, etc. Inconvenient to operate

Method used

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  • Nucleic acid synthesis solid phase carrier, preparation method, nucleic acid synthesis device containing the carrier, and nucleic acid synthesis method
  • Nucleic acid synthesis solid phase carrier, preparation method, nucleic acid synthesis device containing the carrier, and nucleic acid synthesis method
  • Nucleic acid synthesis solid phase carrier, preparation method, nucleic acid synthesis device containing the carrier, and nucleic acid synthesis method

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preparation example Construction

[0035] The embodiment of the present invention provides a method for preparing a nucleic acid synthesis solid phase carrier, comprising the following steps:

[0036] S1, obtaining water-based magnetic fluid. Preferably, the step of obtaining water-based magnetic fluid comprises:

[0037] S11, dissolving ferric chloride hexahydrate and ferric chloride tetrahydrate in a molar ratio of 1 to 3:1 in water to obtain Fe 3+ Iron salt solution with a concentration of 0.2-0.5mol / L. Further, the molar ratio of ferric chloride hexahydrate to ferric dichloride tetrahydrate is 2:1, and the concentration of the iron salt solution is 0.3 mol / L.

[0038] S12, under an inert atmosphere, add a precipitating agent, stir in a water bath at 70-90° C. for 20-40 min, and separate to obtain a precipitated product. Further, nitrogen or helium is selected as the inert gas. Ammonia water is selected as the precipitating agent.

[0039] Further, in a preferred embodiment of the present invention, in ...

Embodiment 1

[0072] A kind of MCPG provided by the present embodiment is prepared according to the following steps:

[0073] (1) Dissolve ferric trichloride hexahydrate and ferric trichloride tetrahydrate in water at a molar ratio of 2:1 and add them to a three-necked flask so that the concentration of iron ions in the solution is 0.3mol / L. Under the protection of nitrogen, add excess ammonia water to form a black precipitate. At the same time, stir vigorously, heat in a constant temperature water bath at 80°C, and maintain PH = 10-11, and stop stirring after 30 minutes. Suction filtration, washing with deionized water until the filtrate flowing down is neutral, and the precipitated product is obtained.

[0074] (2) Disperse the obtained precipitated product in 50ml of water, add 1ml of tartaric acid, heat and stir in a constant temperature water bath at 70°C, and prepare a water-based magnetic fluid after 30min.

[0075] (3) 100mgFe 3 o 4 The fine particles are dispersed in a system of...

Embodiment 2

[0081] A nucleic acid synthesis method provided in this embodiment comprises the following steps:

[0082] (1) Take by weighing 0.5mgMCPG (provided in embodiment 1), insert the electromagnetic rod (hereinafter referred to as reaction rod) of adsorption MCPG in the dichloromethane solution of 1% trifluoroacetic acid, power on and off 30 times, make MCPG in the reaction solution Repeatedly disperse, aggregate, and react to remove the 5' hydroxyl protecting group DMT.

[0083] (2) Mix the required nucleotide monomers and tetrazolium to obtain an activated nucleoside phosphorous acid intermediate, insert the reaction rod and turn it on and off 50 times.

[0084] (3) Insert the reaction rod into the mixed solution of acetic anhydride and acetonitrile and turn it on and off 50 times;

[0085] (4) Insert the reaction rod into the mixture of iodine, pyridine and water and turn it on and off 50 times

[0086] (5) Steps (1)-(4) are repeated until the desired nucleic acid sequence is o...

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Abstract

The invention provides a solid-phase carrier for nucleic acid synthesis, the preparation method of which is as follows: prepare a water-based magnetic fluid, add it to a silica sol precursor containing soft magnetic material powder, let it stand until a gel is produced, and then the gel is 600-900° C. heat treatment for 1-3 hours to obtain magnetically controlled pore glass beads MCPG. Activation of MCPG enables the attachment of various nucleotide monomers. The present invention also relates to a nucleic acid synthesis device, which includes the above-mentioned MCPG, an electromagnetic rod formed by embedding an electromagnet in a housing, and a reaction plate. The reaction rod is loaded with an appropriate amount of activated MCPG, and then the reaction rod is inserted into each reaction cell in the reaction plate, and the adsorption and release of MCPG is realized by controlling the power on and off of the electromagnet, and the synthesis of nucleic acid fragments is completed. The device can replace the existing plastic synthesis column, and the scale of nucleic acid synthesis from 1-50000nmol can be realized in the same set of equipment by controlling the amount of magnetically controlled pore glass beads.

Description

technical field [0001] The invention relates to the technical field of nucleic acid synthesis, and in particular to a nucleic acid synthesis solid phase carrier, a preparation method, a nucleic acid synthesis device containing the carrier, and a nucleic acid synthesis method. Background technique [0002] The artificial chemical synthesis of nucleic acid fragments began in the early 1950s. In 1980, after the introduction of a fully automatic solid-phase nucleic acid synthesizer, it became possible to synthesize nucleic acid fragments quickly and efficiently, which greatly promoted the vigorous development of bioengineering technology. In the prior art, DNA is synthesized by the solid-phase phosphoramidite triester method: in the solution, β-acetonitrile phosphoramidite is used to chemically synthesize nucleic acid fragments, and the synthesis of nucleic acid chains is carried out from 3'→5' direction, usually at the 3' end The first base is bound to Controlled Pore Glass (CP...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N15/10C03C14/00C03C4/00C12M1/42C12M1/00
CPCC03C4/00C03C14/00C03C2203/20C12N15/1006C12N15/1013
Inventor 郑志凌陈波朱志强许全贵
Owner 艾维迪亚(深圳)医疗科技有限公司
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