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Recombinant microorganism and method for producing poly-γ-glutamic acid

A technology of recombinant microorganisms and glutamic acid, which is applied in the direction of recombinant DNA technology, bulk chemical production, and the use of vectors to introduce foreign genetic materials, etc., can solve the problems of inefficient production of PGA

Active Publication Date: 2015-11-25
KAO CORP
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0029] However, the above-mentioned non-patent documents 1 to 3, 7 and patent document 1 do not disclose technology for improving the generation of PGA, and there is a problem that PGA cannot be efficiently generated in the existing knowledge.

Method used

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  • Recombinant microorganism and method for producing poly-γ-glutamic acid
  • Recombinant microorganism and method for producing poly-γ-glutamic acid
  • Recombinant microorganism and method for producing poly-γ-glutamic acid

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Embodiment

[0094] Hereinafter, the present invention will be described in further detail using examples, but the technical scope of the present invention is not limited to the following examples. Table 1 shows the names of the primers used in this example, their sequences, and their serial numbers. In addition, in the nucleotide sequences shown in Table 1, the underlined part indicates the added restriction enzyme recognition sequence.

[0095] Table 1

[0096] Primer

manufacture example 1

[0097] [Manufacturing example 1] Construction of carrier (1)

[0098] This production example shows the cloning method of the gene group involved in PGA synthesis (refer to figure 1 ).

[0099] First, the Bacillus The chromosomal DNA prepared by sp.KSM-366 strain (FERMBP-6262) was used as a template, and the primer pair of pgsB-F (sequence number 12) and pgsA-R (sequence number 13) shown in Table 1 was used to prepare pgsBCA A 2.9 kb DNA fragment of the gene (A). Additionally, serial number 47 indicates Bacillus sp.KSM-366 strain pGS The base sequence of the gene, the sequence number 48 indicates the above pGS The amino acid sequence of the protein encoded by the gene, represented by SEQ ID NO: 49 pGS The base sequence of the gene, the sequence number 50 indicates the above pGS The amino acid sequence of the protein encoded by the gene, represented by SEQ ID NO: 51 pGS The base sequence of the gene, the sequence number 52 indicates the above pGS The amino ac...

manufacture example 2

[0103] [Manufacturing example 2] Construction of carrier (2)

[0104] This production example shows the cloning method of the gene group involved in PGA synthesis (refer to image 3 ).

[0105] First, the Bacillus subtilis The chromosomal DNA prepared by Marburg No.168 strain (Bacillus subtilis strain 168) was used as a template, and the primer pair of P_spoVGFW2 (SEQ ID NO: 24) and P_spoVG / CmR (SEQ ID NO: 25) shown in Table 1 was used to prepare a DNA fragment of 0.9 kb on the upstream side (spoVG-UP) for use in spoVG The chloramphenicol resistance gene ( cm r ). Similarly, use the primer pair of P_spoVG / CmF and (SEQ ID NO: 26) P_spoVGRV (SEQ ID NO: 27) shown in Table 1 to prepare a DNA fragment (spoVG-DW) of 0.6 kb on the downstream side (the primers of P_spoVGRV are designed as Will spoVGThe start codon gtg of the gene ORF was changed to atg). A 0.9 kb DNA fragment (comp_Cm) of the chloramphenicol resistance gene was prepared using the plasmid pC194 as a template ...

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Abstract

A recombinant microorganism having poly-³-glutamic acid-producing ability, prepared by introducing a Bacillus subtilis pgsB gene or a gene functionally equivalent thereto, and a Bacillus subtilis pgsC gene or a gene functionally equivalent thereto, among a group of genes relating to synthesis of poly-³-glutamic acid, into a host microorganism, in which no Bacillus subtilis pgsA gene or a gene functionally equivalent thereto is being introduced into the host microorganism; and a method of producing poly-³-glutamic acid employing the recombinant microorganism.

Description

[0001] technology area [0002] The present invention relates to a recombinant microorganism capable of producing poly-γ-glutamic acid, and a method for producing poly-γ-glutamic acid using the recombinant microorganism. Background technique [0003] Poly-γ-glutamic acid is a polymer compound in which the γ-position carboxyl group and the α-position amino group of glutamic acid are linked by peptide bonds. Poly-gamma-glutamic acid is also sometimes referred to as gamma-polyglutamic acid. Poly-γ-glutamic acid as Bacillus natto ( Bacillus subtilis var. natto ) is known, and in recent years, it has attracted attention as a new type of polymer material due to its various properties. In the description below, poly-γ-glutamic acid is abbreviated as PGA. [0004] As microorganisms producing PGA, some bacillus ( Bacillus ) of the genus Bacteria and its close relatives ( Bacillus subtilis var. chungkookjang , Bacillus icheniformis , Bacillus megaterium , Bacillus anthra...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N1/21C12N15/09C12P13/00
CPCC12N15/74C12N9/93C12P21/02Y02P20/52
Inventor 泽田和久萩原浩
Owner KAO CORP
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