cDNA overall length nucleotide sequence of human EBLN-1 gene and cloning method thereof

A technology of EBLN-1 and nucleotide sequence, which is applied in DNA preparation, recombinant DNA technology, genetic engineering, etc., can solve the problems of lagging EBLN-1 physiological function and pathological effect research, full-length cDNA sequence is not clear, etc.

Inactive Publication Date: 2013-06-19
CHONGQING MEDICAL UNIVERSITY
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Problems solved by technology

[0005] Commonly used research strategies for new genes mainly include bioinformatics and in vivo expression analysis of new genes, research on gain-of-function and inactivation strategies, and research on interacting proteins of gene-encoded products, all of which depend on the structural model of the gene. So far, the full-length cDNA sequence of the EBLN-1 gene is still unclear, which seriously lags behind the subsequent research on the physiological function and pathological effects of EBLN-1

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  • cDNA overall length nucleotide sequence of human EBLN-1 gene and cloning method thereof
  • cDNA overall length nucleotide sequence of human EBLN-1 gene and cloning method thereof
  • cDNA overall length nucleotide sequence of human EBLN-1 gene and cloning method thereof

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[0031] The content of the present invention will be described in more detail below in conjunction with the embodiments and the accompanying drawings, but it should not be understood that the scope of the present invention is limited to the following embodiments. Without departing from the above-mentioned technical idea of ​​the present invention, various replacements and changes made according to common technical knowledge and customary means in this field shall be included in the scope of the present invention.

[0032] 1. Extraction of total RNA from human oligodendroglioma cell lines

[0033] Total RNA was extracted from human oligodendroglioma cell lines (purchased from ATCC cell bank, USA) by Trizol method. Take a cell sample stored at -80°C, add 1 ml Trizol, shake vigorously for 30 seconds to ensure complete dissolution, place at room temperature for 10 minutes, and centrifuge at 4°C for 10 minutes. Add 200ul of chloroform, mix by inversion for 15 seconds, centrifuge at...

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Abstract

The invention discloses a cDNA overall length nucleotide sequence of a human EBLN-1 gene and a cloning method thereof. According to the invention, the cloning method comprises the following steps of: respectively amplifying a 5' end fragment and a 3' end fragment of human EBLN-1 gene according to 5'RACE and 3'RACE technologies and sequencing; then, splicing to obtain the cDNA overall length nucleotide sequence of the human EBLN-1 gene according to the 3' end sequence, the 5' end sequence and a known cDNA part sequence; performing analysis of an opened reading frame and an untranslated region; and deducing to obtain an amino acid sequence which codes the EBLN-1 protein according to the opened reading frame. The cDNA overall length nucleotide sequence of the human EBLN-1 gene can be used for subsequently expressing a recombinant protein for preparing corresponding antibodies or performing structural biological study, for studying EBLN-1 gene function through RNA interference and overexpression or further studying the effect of the untranslated region in regulation of gene expression, and finally laying a foundation for illustrating the physiological action of EBLN-1 gene in human body and the relation with diseases, in particular mental diseases.

Description

technical field [0001] The invention belongs to the field of biotechnology, and more specifically relates to a human endogenous Borna-like N element-1 (EBLN-1) isolated from human oligodendroglioma cells The full-length cDNA sequence of the gene and its cloning method. Background technique [0002] Borna disease virus (BDV) is a non-segmented single-stranded negative-sense RNA virus first discovered in Germany in 1885. It is strictly neurotropic. Its genome encodes 6 viral proteins, nucleoprotein ( N) is the main infection marker and pathogenic protein. In 1996, German scientists successfully isolated BDV from patients with depression, proving that BDV infection also exists in humans (Bode L, et al. Mol Psychiatry. 1996). In recent years, a large number of epidemiological investigations at home and abroad have suggested that there may be a correlation between the occurrence of human mental diseases, including depression, and BDV infection. A concept was proposed. [0003...

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/12C07K14/47C12N15/10
Inventor 谢鹏张亮杨德雨雷阳黄荣忠
Owner CHONGQING MEDICAL UNIVERSITY
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