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Transgene zebra fish system with abcb4 and establishing method thereof

A method for establishing a zebrafish line, which is applied in the field of genetic engineering, can solve problems such as the incomplete understanding of the role and the lack of establishment of a transgenic zebrafish animal model, and achieve the effect of a long animal growth cycle

Active Publication Date: 2016-11-09
GUIZHOU MEDICAL UNIV
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Problems solved by technology

Therefore, the promoter of the zebrafish abcd4 gene, its prediction and cloning methods, and the role of the abcb5 gene in the mechanism of multidrug resistance in zebrafish are not yet fully understood.
Moreover, the zebrafish abcd4 gene promoter and its prediction and cloning methods based on multidrug resistance research have not yet established a transgenic zebrafish animal model

Method used

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  • Transgene zebra fish system with abcb4 and establishing method thereof
  • Transgene zebra fish system with abcb4 and establishing method thereof
  • Transgene zebra fish system with abcb4 and establishing method thereof

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Effect test

Embodiment 1

[0053] 1 zebrafish abcb4 Gene promoter prediction and cloning

[0054] 1.1 Zebrafish abcb4 Gene promoter prediction

[0055] According to the NCBI website, through bioinformatics analysis, the zebrafish abcb4 The gene sequence of about 8000 bp between the gene transcription start site and the upstream adam15 gene was used as a template, and the promoter range was predicted by the Promoter 2.0 Prediction Server promoter prediction website.

[0056] 1.2 Zebrafish abcb4 Gene promoter cloning

[0057] (1) Select about 20-30 5dpf zebrafish embryos, put them into RNAse-Free 1.5ml Epp tubes, and wash the embryos with 1×PBS solution;

[0058] (2) After cleaning, add 500μl Lysis Buffer (which contains proteinase K, the final concentration is 0.17mg / ml), seal and digest in a water bath at 55°C for 1 hour;

[0059] (3) Centrifuge at 14,000rpm at room temperature for 10 minutes, and transfer the supernatant to a new RNAse-Free 1.5ml Epp tube;

[0060] (4) Add 500 μl of phenol / c...

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Abstract

The invention discloses a transgene zebra fish system and an establishing method thereof. The transgene zebra fish system is characterized in that at the premises of not changing the expression of gene abcd4 of zebra fish, the change of expression of gene abcd4 is monitored by EGFP (enhanced green fluorescent protein), namely that the transgene zebra fish system is an EGFP-expressed Tg (abcd4: EGFP) transgene zebra fish system which is driven by an abcd4 gene promoter. The transgene zebra fish system has the advantages that at the premises of not changing the expression of gene abcb4 (which is homologous with the gene ABCB1 of human body) of the zebra fish, the change of expression of gene abcd4 is monitored by EGFP, namely that the transgene zebra fish system is the EGFP-expressed Tg (abcd4: EGFP) transgene zebra fish system which is driven by the abcd4 gene promoter; the early experiment basis is provided for gene abcb 4 in the study of multi-drug resistant mechanism.

Description

technical field [0001] The invention relates to a transgenic zebrafish line and a method for establishing the same, belonging to the technical field of genetic engineering. Background technique [0002] Multidrug resistance (multidrug-resistance, MDR) is a common reason for the failure of tumor chemotherapy, but for now, chemotherapy is still the main method of cancer treatment. Therefore, in judging the prognosis and survival rate of cancer, the occurrence of multidrug resistance has become an important indicator. The definition of multidrug resistance refers to the simultaneous resistance of tumors to multiple drugs with different structures and pharmacology. There are many mechanisms for the generation of multidrug resistance. The traditional view is that it is caused by gene mutations in certain cells in the tumor. However, some pathophysiological factors, such as hypoxia, regulating oncogenes, tumor suppressor genes, cell Changes in apoptosis factors may also promote ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A01K67/027C12N15/89C12N15/66C12N15/65
CPCA01K67/0275A01K2227/40A01K2267/0393C12N15/65C12N15/66C12N15/89
Inventor 舒莉萍何志旭孙琮杰胡荣英金璐何思佳吴西军
Owner GUIZHOU MEDICAL UNIV
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