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Tumor-specific gene expression cassette, recombinant expression vector, construction method and application

A gene expression cassette, tumor-specific technology, applied in the field of tumor treatment, can solve the problems of inefficient expression and low gene expression efficiency.

Active Publication Date: 2021-06-15
XINXIANG MEDICAL UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

However, when mda-7 is used in combination with other genes, there is a problem of low gene expression efficiency, and it cannot be expressed efficiently to play a role

Method used

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  • Tumor-specific gene expression cassette, recombinant expression vector, construction method and application
  • Tumor-specific gene expression cassette, recombinant expression vector, construction method and application
  • Tumor-specific gene expression cassette, recombinant expression vector, construction method and application

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preparation example Construction

[0034] The preparation method of the recombinant expression vector of an embodiment of the present invention, it comprises the following steps:

[0035] Provide an expression vector with a promoter and a terminator, respectively insert the target gene, linker, first suicide gene and second suicide gene into the expression vector by double enzyme digestion and ligation, and obtain a structure containing a promoter-target gene-linker - A recombinant expression vector of the first suicide gene-second suicide gene-terminator. Wherein, the target gene is the melanoma differentiation-related gene 7, the linker is the connecting peptide GS gene sequence, the first suicide gene is the herpes simplex virus thymidine kinase gene, and the second suicide gene is the cytosine deaminase gene.

[0036] In a specific example, the expression vector is a lentiviral vector pLenti-CMV-MCS-3FLAG, the structure of which is as follows figure 1 shown.

[0037] Melanoma differentiation-related gene ...

Embodiment 1

[0041] Synthetic primers were designed according to the mda-7 sequence reported by GenBank (No: U16261.1, 621bp), and the gene fragment was amplified by PCR. Using Escherichia coli E.coli BL21 genomic DNA as a template, synthetic primers were designed according to the sequence of the cytosine deaminase gene reported by GenBank (NO: AY552602, 1284bp), and the gene fragment was amplified by PCR. Using the plasmid pHSV-TK as a template, synthetic primers were designed according to the herpes simplex virus thymidine kinase gene sequence (NO: X03764, 1131bp) reported by GenBank, and the gene fragment was amplified by PCR. Select a structure such as figure 1 The shown lentiviral vector pLenti-CMV-MCS-3FLAG is used as an empty vector, and the linker is the connecting peptide GS gene sequence (GGGGS) 3, and the sequence of the connecting peptide GS is shown in SEQ ID NO.1. Melanoma differentiation-related gene 7, connecting peptide GS gene sequence, herpes simplex virus thymidine kin...

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Abstract

The invention relates to a tumor-specific gene expression box, a recombinant expression vector, a construction method and application. Including a promoter, a target gene, a linker, a first suicide gene, a second suicide gene and a terminator connected in sequence; wherein, the target gene is melanoma differentiation-related gene 7, and the linker is a connecting peptide GS gene sequence, the The first suicide gene is herpes simplex virus thymidine kinase gene, and the second suicide gene is cytosine deaminase gene. The present invention not only combines mda-7 with extremely strong anti-tumor ability and double suicide fusion gene TK-CD, enhances anti-tumor activity, can effectively identify tumor sites, but also connects them by linking peptide GS gene sequence, so that The tumor-specific gene expression cassette or the recombinant expression vector containing the tumor-specific gene expression cassette has high transcriptional activity and can be expressed efficiently, providing a new way for tumor treatment.

Description

technical field [0001] The invention relates to the field of tumor treatment, in particular to a tumor-specific gene expression cassette, recombinant expression vector, construction method and application. Background technique [0002] A large number of studies in recent years have confirmed that the biological basis of tumor occurrence and development is gene mutation. Therefore, tumor gene therapy has become a hot spot in oncology research. In gene therapy, how to combine tumor-related genes to overcome their respective defects, enhance the anti-tumor effect, and specifically and efficiently express in tumor cells or tumor tissues is of great significance to the efficiency and safety of gene therapy. , is also one of the challenges encountered in gene therapy. [0003] Melanoma differentiation-associated gene-7 / interleukin-24 (mda-7 / IL-24) is a tumor-inhibiting cytokine belonging to the IL-10 family discovered by subtractive hybridization, which can specifically It can ...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N15/63C12N15/66A61K48/00A61P35/00
CPCA61K48/005A61P35/00C07K14/4748C12N15/63C12N15/66
Inventor 杨瑞张哲莹胡育菡徐志浩张艳芬
Owner XINXIANG MEDICAL UNIV
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