Gene expression component, constructed cloning vector and application

Abstract

The invention provides a gene expression component, a constructed cloning vector and application. The gene expression component encodes beta-galactosidase alpha peptide and omega peptide; the gene expression component includes a constitutive promoter, and the constitutive promoter promotes an expressed LacZ[alpha] gene and LacZ[omega] gene. The constitutive promoter is used to regulate the expression of the LacZ[alpha] gene and LacZ[omega] gene, a constructed pWizard plasmid can produce substances with beta-galactosidase activity in a host cell without IPTG induction, the application scope ofthe host cell is expanded, the pWizard plasmid is a high-copy plasmid, therefore, the copy number of the LacZ[alpha] gene and the LacZ[omega] gene in a host bacteria is high, the expression amount ofthe host cell to an enzyme with beta-galactosidase activity is significantly increased, the color development effect and the color development speed are enhanced, and the color development time is shortened.

Description

technical field [0001] The invention belongs to the technical fields of molecular biology and genetic engineering, and relates to a gene expression component and its constructed cloning vector and application, in particular to a gene expression component encoding β-galactosidase α peptide and ω peptide, and the constructed Cloning vector and its construction method and application. Background technique [0002] PCR is an important technique in the fields of molecular biology and genetic engineering, and experimenters often need to clone PCR products into vectors. Currently commonly used cloning vectors are mainly based on the principle of blue-white screening. The blue-white screening method brings great convenience for picking positive clones. The vector that can be used for blue-white screening contains a lacZ' gene, which can encode an α-peptide chain, and contains a multiple cloning site (MCS) for cloning foreign DNA in the lacZ' gene. Existing vectors with blue-white ...

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Problems solved by technology

[0003] However, the existing vectors with blue-white screening function, such as the widely used pUC series vectors, have the following disadvantages: First, the scope of application of strains is limited, and this vector is only suitable for β-galactosidase-deficient Genetically engineered bacteria, but Escherichia coli without mutations in β-galactosidase are often used in practical applications, such as STBL2 and STBL3 strains, which cannot use the blue-white phenotype to screen positive clones; second, each Escherichia coli contains a lower Genome copy number leads to low expression of omega peptide. When the strain is transformed into a plasmid expressing α peptide, the content of β-galactosidase is low, and it takes a long time for the strain to form obvious blue spots. Low, affecting production efficiency and large-scale industrial application; third, this carrier needs to use a plate containing IPTG, but IPTG is expensive and toxic to the human body, with high cost and low safety

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