Establishment of self-activated Gal4/UAS system expression cassette capable of improving gene expression

Abstract

The invention discloses establishment of a self-activated Gal4/UAS system expression cassette capable of improving gene expression. The expression cassette with good improvement effect is obtained byoptimizing the number of UAS and the types of GAL 4 activation/binding domains; the expression cassette is started by the strong promoter CMV of the expression cassette and improves transcription andtranslation efficiency in virtue of a Gal4/UAS system, so the effects of automatic activation and cascade amplification are obtained; high improvement effect is maintained in 5 to 10 days after transfection, and highest expression activity is obtained in the eighth day, so it is proved that the expression cassette can improve the stability of gene expression; and improvement effect is about 9 times better in CHO cells specially producing recombinant proteins. Compared with conventional gene expression cassettes, the expression cassette provided by the invention is simple in structure and substantial in improvement effect, and provides an approach and novel method for non-virus transfection approaches of gene therapy and enhancement of the expression of target genes in production of recombinant proteins.

Description

technical field [0001] The invention relates to the establishment of a self-activating Gal4 / UAS system expression box for enhancing gene expression, and belongs to the field of biotechnology. Background technique [0002] The Gal4 / UAS system was first discovered in Saccharomyces cerevisiae (Oshima Y et al.1984), and it consists of two parts: the gene encoding the yeast transcription activator GAL4 (Transcription Factor GAL4) and the upstream activation sequence (Upstream Activation Sequence, UAS , Giniger E and Ptashne 1987; Giniger Edward et al. 1985). Saccharomyces cerevisiae GAL4 protein consists of 881 amino acid residues, mainly including two parts: DNA binding domain and DNA activation domain (Laughon et al.1984; Oshima Yasuji 1982). The DNA binding domain is located at the 1-65 peptide segment at the N-terminal of the protein, and the 7-Cys-X2-Cys-X6-Cys-X6-Cys-X2-Cys-X6-Cys40 peptide segment in this domain constitutes a Zn2Cys6 structure, which binds DNA sites (Hel...

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Problems solved by technology

[0006] At present, there are no reports on the expression of target genes in humanized cells using Gal4 / UAS-based expression systems, and the existing expression systems still have low expression levels of target genes, short expression times, and poor stability of enhanced expression, etc. Insufficient, which is not conducive to its development and application in non-viral transfection pathways of gene therapy and DNA vaccines

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