Method for detecting compound genotoxicity by biomarker and use of the biomarker

A technology of biomarkers and genotoxicity, which is applied in the field of using biomarkers to detect the genotoxicity and application of compounds, can solve the problems of quantitative innovation and research depth gap, and achieve the effect of simple operation, strong sensitivity and high sensitivity

Inactive Publication Date: 2012-10-17
SHANGHAI INST OF MATERIA MEDICA CHINESE ACAD OF SCI
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  • Application Information

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Problems solved by technology

At present, the Human Genome Project has entered the stage of functional genomics research. The rapid development of gene function research has brought opportunities for the exploration and research of new molecular biomarkers. my country has also made some achievements in this field, but in terms of quantity There is a big gap with foreign countries in terms of quality, innovation and research depth

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  • Method for detecting compound genotoxicity by biomarker and use of the biomarker
  • Method for detecting compound genotoxicity by biomarker and use of the biomarker
  • Method for detecting compound genotoxicity by biomarker and use of the biomarker

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Experimental program
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Embodiment 1

[0033] In the research of exploring molecular biomarkers, microarray technology has played a role that cannot be underestimated. Through gene expression profile and genome sequence analysis, combined with other detection methods such as Real Time-PCR to confirm mRNA expression and Western Blot to detect protein expression levels, the effector molecular biomarkers that cause toxicity can be finally determined. We selected 7 known genotoxic compounds ENU, DPN, DEN, DMN, o-AAT, DBP and DMBA with different mechanisms of action, and used the doses that can produce genotoxicity in the preliminary experiment on 8-week-old normal males B6C3F1 mice (provided by Japan CLEA Company) were given a single intraperitoneal administration, and three toxic but not genotoxic compounds DEHP, PB and EtOH were selected, and the mice were given a single dose of 1 / 2 half lethal dose (LD50) Administration, Table 1 shows the above compounds and the corresponding doses. Liver total RNA was extracted 4 ...

Embodiment 2

[0038] (1) In vivo Real Time-PCR experiment to verify the expression of BC gene

[0039] We selected 4 genotoxic substances ENU, DPN, DEN, o-AAT and 2 non-genotoxic substances DEHP and EtOH investigated in the gene chip experiment to verify the reproducibility of the chip results and the validity of the markers. Normal male C57 mice weighing 20-22 g were used, provided by the Shanghai Experimental Animal Center of the Chinese Academy of Sciences. Animals were kept in SPF grade animal rooms, and animal experiments were carried out after at least 3 days of adaptation. Set up ENU, DPN, DEN, o-AAT, DEHP, and EtOH administration groups and control groups, with 4 mice in each group, and use the same dose as the gene chip test to administer a single intraperitoneal administration to the mice, 0.2ml / 10g body weight, and the control group was given normal saline. Liver RNA was extracted 4h and 20h after administration, and Real Time-PCR was performed to investigate the expression of ...

Embodiment 3

[0055] like figure 1 As shown, the test substance is administered to mice at different doses, or the cultured cells derived from mice are treated, and the vehicle of the test substance is used as a control group to administer the mice or cells. After a certain period of time after administration, the total RNA of mouse tissues or cells was extracted, and cDNA was obtained by reverse transcription, and cDNA was used as a template to detect the expression of BC genes in specific tissues or cells. The specific implementation method is as follows.

[0056] (1) Extraction of total RNA from tissues or cells

[0057] Preservation of tissues: After the mice were sacrificed, the tissues such as heart, liver, spleen, lung, and kidney were quickly separated, submerged in a centrifuge tube containing RNAlater (at least 1ml of RNAlater was required for each 100mg of tissue), and after standing overnight at 4°C, If stored at -20°C or -80°C, the RNA can remain intact for a long time withou...

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Abstract

The invention discloses a method for detecting compound genotoxicity by a biomarker and a use of the biomarker. The biomarker is a mice BC gene. The method comprises the following steps of 1, treating a mice or cells by a compound to obtain mice tissue or a cell sample, and 2, detecting the expression of a biomarker in the mice tissue or the cell sample, wherein the biomarker is a mice BC gene and through the expression of the biomarker, genotoxicity of the compound is detected. The method for detecting compound genotoxicity has the characteristics of accuracy, high sensitivity and easy operation, can be used as a candidate method for evaluating drug safety and can be used for early screening of genotoxicity of a new drug.

Description

technical field [0001] The invention relates to a method for detecting compound genotoxicity using biomarkers and the application of the method in early genotoxicity screening of new drug development. Background technique [0002] Since the 20th century, with the advancement of science and technology, the improvement of the level of industrialization, and the expansion of human activity areas, new chemical substances in the human living environment have continued to increase. Among them, some genotoxic substances can directly damage the genetic material of organisms and cause Changes in genes and chromosomes can induce cancer, and even pass mutations to offspring through germ cells. The prediction and evaluation of the genotoxicity of these chemical substances has become an important research content of environmental science, and the development and improvement of genotoxicity detection and evaluation technology is of great and far-reaching significance for the protection of...

Claims

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Application Information

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IPC IPC(8): C12Q1/68
Inventor 栾洋任进武元峰王以政戚新明
Owner SHANGHAI INST OF MATERIA MEDICA CHINESE ACAD OF SCI
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