Preparation of CRISPR (clustered regularly interspaced short palindromic repeats)/Cas9 nano gene system and application of CRISPR/Cas9 nano gene system in transfection

A gene system and nanotechnology, applied in the direction of genetic engineering, biochemical equipment and methods, and other methods of inserting foreign genetic materials, can solve the problems of difficult quality control of transfection vectors, low efficiency, and ineffective transfection effects , to achieve the effect of improving transfection efficiency and simple operation

Inactive Publication Date: 2018-04-13
FUZHOU UNIV
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Problems solved by technology

[0007] The purpose of the present invention is to provide a preparation of a novel CRISPR/Cas9 nano-gene system and its application in transfection, so as to s...

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  • Preparation of CRISPR (clustered regularly interspaced short palindromic repeats)/Cas9 nano gene system and application of CRISPR/Cas9 nano gene system in transfection
  • Preparation of CRISPR (clustered regularly interspaced short palindromic repeats)/Cas9 nano gene system and application of CRISPR/Cas9 nano gene system in transfection
  • Preparation of CRISPR (clustered regularly interspaced short palindromic repeats)/Cas9 nano gene system and application of CRISPR/Cas9 nano gene system in transfection

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[0039] In order to make the content of the present invention easier to understand, the technical solutions of the present invention will be further described below in conjunction with specific embodiments, but the present invention is not limited thereto.

[0040] 1. Preparation of CRISPR@CS

[0041] At room temperature, configure a 0.1% (v / v) acetic acid solution for use, weigh 2g of chitosan (CS) with an electronic balance and add it to the acetic acid solution, slowly add the acetic acid dropwise until the chitosan is completely dissolved in the solution, and magnetically After stirring evenly, a 2% (w / v) chitosan / acetic acid solution was obtained. After adding a certain amount of CRISPR, incubated at 150 rpm for 10 min at 20°C, the mass ratio of chitosan to CRISPR (w / w) was 1:1, 1:2, 1:3, 1:4, 1:5, and five CRISPR@CS nanocomposites with different mass ratios were prepared.

[0042] 2. Particle size and potential analysis and detection of CRISPR@CS nanocomposites

[0043...

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Abstract

The invention belongs to the technical field of biology and particularly relates to preparation and cell transfection application of a CS-based (chitosan-based) and CRISPR/Cas9 plasmid-constructed nano gene drug. The preparation comprises dissolving chitosan into acetic acid solution, dropwise adding the chitosan acetic acid solution into water, adding in CRISPR in mass proportion to the chitosan,encapsulating and carrying CRISPR/Cas9 with the chitosan to prepare a novel gene drug nano particle for good gene transfection effects or targeted exogenous gene introduction and further to achieve the effects of transferring gene drugs into cells. The novel nano compound of CRISPR@CS structured on the CS and the CRIPPR through an optimal feeding ratio can significantly improve the transfection efficiency of genes in cells to convey more genes into the cells to take effects, thereby being expected to being widely applied in the field of gene treatment through gene drug delivery.

Description

technical field [0001] The invention belongs to the field of biotechnology, and in particular relates to the preparation and cell transfection application of a nano-gene drug constructed on the basis of chitosan (CS) and CRISPR / Cas9 plasmid. Background technique [0002] Chitin exists in a very high amount in nature and is rich in sources. It is a biological polysaccharide second only to cellulose on the earth. It is widely found in animals, plants, and various fungi. Studies have found that especially crustaceans (crabs, shrimps, etc.) ) content is the highest, up to 58%~85%, followed by Insecta, etc., whose content can reach 20%~60%; Also contains a certain amount of chitin. Chitosan is a derivative obtained after the N-acetylamino group on the sugar group in chitin is removed. Generally speaking, chitosan is a straight chain connected by β-(1,4) glycosidic bonds. For each unit, more than 55% of the N-acetylamino group can be called chitosan, or in other words, the amoun...

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Application Information

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IPC IPC(8): C12N15/90C12N9/22
CPCC12N9/22C12N15/907
Inventor 邵敬伟吴鹏宇沈志春张冰晨
Owner FUZHOU UNIV
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