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Interleukin-2 mutation gene and its preparation

A technology for mutant gene and interleukin, which is applied in the field of interleukin-2 mutant gene and its preparation, and can solve the problems of high synthesis cost, lack of emphasis, and many mutation sites.

Inactive Publication Date: 2004-10-20
SHENYANG INST OF APPLIED ECOLOGY - CHINESE ACAD OF SCI
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

D.P.Williams et al. synthesized seven segments of nucleotides according to this principle, then connected them, and expressed them in Escherichia coli (D.P.Williams, D.Rcgier, D.Akiyoshi, F.Cenbauffe and J.Murphy.Desin, snthesis and expression of a human interleukin-2gene incorporating the codon usage bias found in highly expressed Escherichia coli gene. Nucleic Acids Research, 1988, 16(22): 10453-10467); this is about 70 bases per segment, and the synthesis cost is also high; in addition There are too many mutation sites, the focus is not prominent, and it is chemically synthesized

Method used

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  • Interleukin-2 mutation gene and its preparation
  • Interleukin-2 mutation gene and its preparation
  • Interleukin-2 mutation gene and its preparation

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Embodiment 1

[0029] An interleukin-2 mutant gene, having the sequence shown in SEQ ID NO: 1:

[0030] (1) Information of SEQ ID NO.1 (see sequence listing)

[0031] (a) Sequence features:

[0032] * Length: 399 bp

[0033] *Type: nucleic acid

[0034] * Chain type: double chain

[0035] *Topology: Linear

[0036] (b) Molecular type: cDNA

[0037] (c) Assumption: No

[0038] (d) Antisense: no

[0039] (e) Original source: interleukin-2 (human interleukin 2)

[0040] Material:

[0041] Bacterial strains and E.coli DH5α, JM109 (DE3) are commercially available;

[0042] Plasmid: a plasmid containing the IL-2 gene obtained by the mutation, a PTE-30a-(+) expression vector, and a 7ZTS vector (see Chinese patent application 02109681.3) constructed in this example (commercially available 7Z can also be used instead);

[0043] Molecular biology reagents: Taq DNA polymerase, Pfu DNA polymerase, dNTP, hind III, Nde I, T4 DNA Ligase are all products of Promega Company in the United States, ge...

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Abstract

An interleukin-2 (IL-2) mutation gene with the base sequence shown by SEQ ID No.1 is prepared from natural IL-2 gene through overlapped PCR for targeted mutation to the base on the gene. Its advantages are less mutant site, high protein expression level, and no change to amino acid sequence of natural protein.

Description

technical field [0001] The invention relates to gene recombination technology, in particular to an interleukin-2 (IL-2) mutant gene and its preparation. Background technique [0002] IL-2 is a kind of lymphokine that activates T-cells to produce and acts between lymphocytes; human IL-2 is a polypeptide chain consisting of 133 amino acid residues (Rene Devos, GeertPlaetinck, Hilde Cheroutre, Guus Simons, Wim Degrave, Jan Tavernier, Erik Remaut and Waltr Fiers, Molecular cloning of human interleukin 2 cDNA and its expression in E. coli., Nucleic Acids Research, 1983, 11(13):4307-4323). In recent years, the study of IL-2 for tumor therapy has made people pay attention to its artificial preparation. It is a very common method to express proteins in Escherichia coli, but there is a problem of rare codons in Escherichia coli when expressing eukaryotic genes; in this way, it becomes a method to mutate rare codons into common codons of Escherichia coli, different The expression ef...

Claims

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Application Information

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IPC IPC(8): C12N15/26C12P19/34
Inventor 吴文芳杨立泉吕安国时成波
Owner SHENYANG INST OF APPLIED ECOLOGY - CHINESE ACAD OF SCI
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