Method for constructing and applying cytochrome C gene-carried slow virus expression plasmid

Abstract

The invention discloses a method for constructing and applying a cytochrome C gene-carried slow virus expression plasmid. The method comprises the following steps: inserting cytochrome C genes into Not I and BamH I multiple cloning sites of a pLenti-Mcherry-Puro carrier; using a constructed pLenti-cytochrome C-Emcherry-Puro recombinant plasmid for transfecting MDA-MB-453 cells; screening and identifying by using puromycin, thereby acquiring a cell strain for stably transfecting cytochrome C gene. The recombinant plasmid contains red fluorescence protein mcherry, so that positive cells can be conveniently observed under a fluorescence microscope; labels of puromycin resistance gene and ampicillin resistance gene are carried by the recombinant plasmid; ampicillin or puromycin can be added for screening during the process of prokaryotic expression or eukaryotic expression; the plasmid establishes a firm basis for further researching biological characteristics and function of cytochrome Cgene and researching and developing anti-tumor drugs.

Description

technical field [0001] The invention belongs to the field of medical molecular biology, and in particular relates to a construction and application method of a lentiviral expression plasmid carrying cytochrome C gene. Background technique [0002] Gene therapy is to introduce the target gene into target cells through gene transfer technology, and use it to regulate the normal function of target cells; compared with traditional treatment methods, gene therapy greatly reduces the pain of patients, especially in the field of tumor gene therapy. Significant advantages have become one of the research hotspots in the field of tumor treatment in recent years. Since the United States approved the world's first gene therapy clinical trial in 1989, a total of 1,384 tumor gene therapy programs have entered the clinical trial stage worldwide. In January 2004, Gendicine, a gene therapy drug independently developed by my country for head and neck tumors, was approved for marketing. It is...

AI Technical Summary

Problems solved by technology

Adenovirus vectors have disadvantages such as short duration of gene expression, repeated administration, easy to produce drug resistance, and small foreign gene fragments; herpes simplex virus vectors have shortcomings such as high immunogenicity