Recombinant dna to control gene expression, the method of monitoring the expression of a target gene, the method of obtaining a target substance

 

The invention relates to biotechnology. The claimed nucleotide sequence to control expression, which controls the expression of a target gene, located after the sequence to control the expression that is dependent on the intracellular concentration of amino acids. In bacteria containing DNA structure from nucleotide sequence to control expression, a promoter located in front of the nucleotide sequence to control the expression and target gene, located after the nucleotide sequence to control expression, the frequency of termination to the nucleotide sequence to control expression at the transcription starting at the specified promoter decreases with increasing intracellular concentrations of amino acids, whereby expression of the target gene is increased. The invention allows to create a system of artificial regulation, providing gain controlled gene expression by increasing the intracellular concentration of regulatory amino acids. 3 S. and 9 C.p. f-crystals, 8 ill., table 1.

Description text in facsimile form (see graphic part)

Faure is then amino acids, containing a region encoding a leader peptide that includes the specified amino acid, and an odd number of segments, not less than 3, and each of the segments or part can form due to the mating grounds mutually exclusive patterns with one of the two adjacent segments, or parts thereof, and the penultimate and last segments or parts thereof are-independent terminator.

2. Recombinant DNA under item 1, characterized in that the codon specified amino acids in the leader peptide is located in the first segment.

3. Recombinant DNA according to any one of paragraphs.1 and 2, characterized in that the leader peptide contains 5 segments.

4. Recombinant DNA according to any one of paragraphs.1-3, characterized in that the nucleotide sequence of each segment or portion, and the nucleotide sequence of the adjacent segment or part represent the inverted repeats.

5. Recombinant DNA according to any one of paragraphs.1-4, characterized in that-independent terminator capable of functioning in bacteria belonging to the genus Escherichia.

6. Recombinant DNA under item 3, characterized in that the first and second segments in order from the 5’-end, and Eona Escherichia Li, the fourth and fifth segments are derived from the sequence of attenuator his-tag operon of Escherichia coli, and the third segment obtained by combining sequences of attenuation tryptophan and his-tag of operons.

7. Recombinant DNA under item 6, wherein the leader peptide contains at least 2 residues of tryptophan.

8. Method of monitoring the expression of a target gene, providing growing in a nutrient medium the bacteria containing recombinant DNA to control gene expression according to any one of paragraphs.1-7, located after the promoter and in front of the target gene, and changes in intracellular concentrations of amino acids that affect gene expression, thus increasing intracellular concentrations of amino acids reduces the frequency of termination on recombinant DNA during the transcription starting at the specified promoter.

9. The method according to p. 8, characterized in that using a bacterium belonging to the genus Eschrichia, Salmonella, or Serratia.

10. The method according to p. 8, wherein the target genes include gene chloramphenicolchloramphenicol (cat), amino acid operons, genes, protein products are involved in the biosynthesis of amino acids, nucleosides and nucleotides, and the genes encoding the alien Besa recombinant DNA according to any one of paragraphs.1-7, located after the promoter and in front of the target gene, and has the ability to products specified substances, and selection of the target substance from the culture fluid.

12. The method according to p. 9, characterized in that the target substances are chloramphenicolchloramphenicol, prokaryotic enzymes, alien proteins, amino acids, nucleotides and nucleosides, vitamins.