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Configuration of cell facter IL-24 eucargon expression carrier and application

A technology of eukaryotic expression vectors and cytokines, applied in the fields of molecular immunology and tumor immunology, can solve the problems of unreported therapeutic effects on liver cancer and small intestine cancer, and biological safety issues, and achieve simple treatment methods and reliable experimental basis Effect

Inactive Publication Date: 2007-06-20
WUHAN UNIV
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Problems solved by technology

[0019] 4.2 There may be an anti-tumor mechanism
[0023] Cloning the IL-24 / MDA-7 gene into a replication-deficient adenovirus to construct the expression vector Ad-mda7 can induce the apoptosis of various tumor cells, but adenovirus as a vector has biological safety problems
And the research on the therapeutic effect of IL-24 on liver cancer and small intestine cancer has not been reported yet.

Method used

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  • Configuration of cell facter IL-24 eucargon expression carrier and application
  • Configuration of cell facter IL-24 eucargon expression carrier and application
  • Configuration of cell facter IL-24 eucargon expression carrier and application

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Embodiment Construction

[0074] 1. Construction method of human IL24 eukaryotic recombinant expression vector

[0075] 1.1 Amplification and purification of the target gene

[0076] Primer design and synthesis: primers were designed according to the human IL-24 GeneBank sequence (NM006850), upstream primer: 5'GC AGATCT AATTTTCAACAGAGGCTG-3', containing Bgl II restriction site; downstream primer: 5'CG GTC GAC TCAGAGCTTGTAGAATTTCTG-3', containing the Sal I restriction site. Primers were synthesized by Shanghai Boya Biotechnology Co., Ltd. Using the plasmid TRAPIL-24 inserted into the complete coding region of human IL-24 as a template, PCR amplification was performed from the second amino acid encoded by the N-terminal of human IL-24 using the above primers. The reaction conditions for the amplification were: pre-denaturation at 95°C for 2 minutes, denaturation at 95°C for 36 seconds, annealing at 56°C for 36 seconds, extension at 72°C for 1 minute and 24 seconds, amplification for 25 cycles, and ex...

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Abstract

A method for configuring the eucaryotic expression carrier of cell factor IL-24 includes designing the ascending and descending primers, PCR amplifying begining from the second amino acid in the code at human IL-24 N terminal by using TRAPIL-24 plasmid as template, cutting off target band, purifying it with reagent kit, configuring and determining recombinant plasmid pEGFP-CI-IL-24, respectively enzyme-severing the PCR product and carrier pEGFP by Bgl II and Sal I, purifying them by reagent kit, linking fragment, culturing, cloning, determining and sequencing. The recombinant cell factor Il-24 can be used to treat liver cancer, small intestine cancer and melanoma.

Description

technical field [0001] The invention belongs to the fields of molecular immunology and tumor immunology, and more specifically relates to a method for constructing a eukaryotic expression vector of cytokine IL-24, and also relates to the use of recombinant cytokine IL-24 expressed by it in the treatment of liver cancer, small intestine cancer and melanoma the use of. Background technique [0002] Interleukin-24 (Interleukin-24, IL-24) is a new cytokine just discovered and determined in 2002 (Caudell EG, Mumm JB, Poindexter N, Ekmekcioglu S, Mhashilar AM, Yang XH, Retter MW, Hill P , Chada S, Grimm EA.The protein product of the tumor suppressor gene, melanoma differentiation-associated gene7, exhibits immunostimulatory activity and is designated IL-24.J Immunol.2002,168(12):6041-6; Zhang Xiaolian, Wang Fubing, new Cytokine IL-24 and its prospect, Journal of Cellular and Molecular Immunology 2003, 19(5), S20-S22.). IL-24 belongs to the IL-10 cytokine family (Wolk K, Kunz S, ...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N15/79C12N15/24C12N15/10A61K48/00A61P35/00
Inventor 章晓联瞿少刚周翔吴建国陈明李平飞
Owner WUHAN UNIV
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