Building method and use of antineoplastic compound screening model

A compound screening and anti-tumor technology, which is applied in biochemical equipment and methods, other methods of inserting foreign genetic materials, measurement/testing of microorganisms, etc., can solve the problem of inconsistency in drug screening at the living level, and achieve improvement Drug screening efficiency, high experimental accuracy, and the effect of improving the probability of becoming a drug

Inactive Publication Date: 2015-05-06
CHANGSHA YINGRUN BIOTECH
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Problems solved by technology

[0003] In the past practice of high-throughput drug screening, drug screening is mainly carried out at the cellular level, and there are great differences between various conditions at the cellular level and the in vivo level. to consistent effect

Method used

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  • Building method and use of antineoplastic compound screening model
  • Building method and use of antineoplastic compound screening model
  • Building method and use of antineoplastic compound screening model

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specific Embodiment 1

[0055] A method for establishing a high-throughput reverse zebrafish SonicHedgehog (Shh) signaling pathway overexpression anti-tumor compound screening model marked by body axis dorsal curvature of the present invention comprises the following steps:

[0056] (1) Synthesis of Morpholino oligonucleotide: the specific sequence of Morpholino oligonucleotide is 5'-TGCAGTATGTTTGTACCGTCGAGTA-3'.

[0057] (2) Zebrafish strain selection: wild-type AB strain was selected as the experimental fish.

[0058] (3) Zebrafish embryo collection, specific steps:

[0059] 3.1) The breeding of zebrafish embryos is carried out in the way of natural pair mating, and 6-10 pairs of adult zebrafish are prepared for each mating, and each pair can produce 200-300 embryos on average.

[0060] 3.2) Collect zebrafish embryos within 0.75 hours after fertilization.

[0061] 3.3) Under the condition of 28.5±0.5°C, incubate the embryos with water for fish culture. The quality of water for fish culture: add 2...

specific Embodiment 2

[0081] The application of a high-throughput reversed zebrafish SonicHedgehog (Shh) signaling pathway overexpression anti-tumor compound screening model marked by body axis dorsal curvature of the present invention can be added to the drug screening group when performing drug screening.

[0082] The specific method of joining the drug screening group is as follows:

[0083] 1) Establish a compound screening model according to the above method.

[0084] 2) Select the appropriate small molecule compound library. In this example, the small molecule compounds were obtained from the compound library obtained from the high-throughput drug screening library of the Institute of Chemical Biology of Vanderbilt University in the United States, and each compound was diluted with DMSO to prepare a 1 mM stock solution.

[0085] 3) Each compound was diluted with embryo culture medium with a final concentration gradient of 5um, 10um, and 20um. At the same time, DMSO at the corresponding conce...

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Abstract

The invention discloses a building method and use of an antineoplastic compound screening model, which can realize high flux screening of an antineoplastic compound for reversion of zebra fish Sonic Hedgehog (Shh) signal path overexpression and is identified by the degree of body axis bending toward the back side, belongs to technical innovation in the technical field of medicine. The antineoplastic compound screening model can be used for preliminary rapid screening of a tumor-inhibition lead compound with latent specificity. A zebra fish tulp3 gene is used as a target gene, and the tulp3 gene is partially silenced by a specific Morpholino oligonucleotide sequence so that Shh path overexpression is realized. When the Shh path is over-expressed, zebra fish shows of body axis specific-bending toward the back side, and the above phenotype is used for identifying Shh path overexpression. Through observation of the zebra fish body appearance, the Shh path overexpression case is directly known. Compared with a cell model, the antineoplastic compound screening model is visual and convenient and is conducive to high flux screening of drugs utilizing Shh path as a target point.

Description

technical field [0001] The present invention relates to the field of biomedicine, in particular to a method for establishing a high-throughput anti-tumor compound screening model for reversing the overexpression of the zebrafish Sonic Hedgehog (Shh) signaling pathway marked by body axis dorsal curvature and its application. A method for establishing a high-throughput drug screening model targeting the Shh signaling pathway. During the drug screening process, it can be determined whether the drug can reverse the overexpression of the Shh signaling pathway by directly observing the phenotype of the dorsal bending of the zebrafish body axis, thereby To achieve the application purpose of drug screening. Background technique [0002] The zebrafish gene is as high as 87% similar to humans, and it has become a good model organism due to its advantages such as small size and convenient reproduction. [0003] In the past practice of high-throughput drug screening, drug screening is ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/89A01K67/027C12Q1/02
Inventor 姜钰超易银沙张万明邹义洲何彰华孙永林刘彩云袁炳秋吴小宁
Owner CHANGSHA YINGRUN BIOTECH
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