Method for exogenous gene expression optimization in salmonella

A Salmonella and exogenous gene technology, applied in the field of optimizing the expression of exogenous genes in Salmonella, to achieve the effect of increasing the expression amount and optimizing the expression

Inactive Publication Date: 2013-05-01
GUANGDONG DAHUANONG ANIMAL HEALTH PRODS +1
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

While Salmonella is an important pathogenic bacterium, there are few reports on the expression of proteins in Salmonella, let alone the optimal expression of genes in Salmonella.

Method used

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  • Method for exogenous gene expression optimization in salmonella
  • Method for exogenous gene expression optimization in salmonella
  • Method for exogenous gene expression optimization in salmonella

Examples

Experimental program
Comparison scheme
Effect test

Embodiment Construction

[0023] Experiment 1: Optimal expression of phage E gene in Salmonella

[0024] 1. First design and synthesize phage E gene-specific primers

[0025] E 1S: ATGGTACGCTGGACTTTGTGGGA 3'

[0026] E 1A: TCACTCCTTCCGCACGTAATTTTTG 3'

[0027] 2. The E gene was obtained by PCR amplification and sequenced, and its sequence was: ATGGT A CGCTGGAC T TTGTGGGATACCCTCGCTTTCCTGCTCCTGTTGAG T TTATTGCTGCCGTC A TTGCT T ATTATGTTCATCCCGTC A AC A TTCAAACG G CC T GTCTC A TC A TGGAAGGCGCTGAATTTACGGAAAACATTATTAATGGCGTCGAGCGTCCGGTTAAA G CCGCTGAATTG T TCGCGTTTACC T TGCGTGTA C GCGCAGGAAAC A CTGACGTTCTTACTGACGCAGAAGAAAAC G TGCGTCAAAAATTA C GTGCG G AAA G GAGTGA.

[0028] 3. Then according to the Salmonella codon usage frequency table, i.e. Table 1 and the amino acid codon table, i.e. Table 2, point mutation is carried out to the E gene, so that its low frequency usage codon becomes a high frequency usage codon, and the E gene mutation The sequence is: ATGGT G CGCTGGAC G TTGTGGGATACCCT...

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Abstract

The present invention provides a method for exogenous gene expression optimization in salmonella. The method comprises the following specific steps: carrying out statistics from a NCBI database to obtain a salmonella codon usage frequency table; analyzing usage frequency of every codon; cloning or synthesizing a target gene, determining a nucleotide sequence of the target gene, adopting a molecular mutation manner or a gene synthesis manner to carry out mutation of the codon having a lowest salmonella codon usage frequency and the codon having a higher salmonella codon usage frequency in the target gene into the codon having a highest usage frequency under a condition of no change of amino acids according to the salmonella codon usage frequency table and an amino acid codon corresponding table to obtain a new nucleotide segment encoding the target gene; adopting a molecular biology method to insert the new nucleotide segment encoding the target gene into an expression vector, transforming the obtained vector to gene engineering escherichia coli, and identifying the obtained clones; and transforming the target gene-containing expression vector into salmonella, wherein the target gene-containing expression vector is identified, and a correct result is obtained.

Description

[0001] technical field [0002] The invention relates to the technical field of gene optimization expression, in particular to a method for optimizing the expression of exogenous genes in Salmonella. [0003] Background technique [0004] Gene cloning and expression are widely used in the production of biologically active substances, and gene expression in heterologous hosts is affected by many aspects, such as promoters, regulatory sequences, terminators, codon usage frequency, etc. In the study of gene expression, researchers pay more attention to the selection of appropriate expression vectors and host systems, but often ignore the substantive issue of whether the gene itself is the best match with the vector and host system. The optimal expression of genes can be achieved by redesigning and synthesizing genes, such as eliminating rare codons and using optimized codons, minimizing secondary structures, adjusting GC content, etc. [0005] There are 64 types of genetic co...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/67C12N15/74
Inventor 徐家华陈瑞爱张东霞汤钦
Owner GUANGDONG DAHUANONG ANIMAL HEALTH PRODS
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