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Novel plasmid autonomously replicable in enterobacteriaceae family

A kind of Enterobacteriaceae, plasmid technology, applied in the field of using the plasmid and microorganisms containing the plasmid

Inactive Publication Date: 2008-02-20
AJINOMOTO CO INC
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

Although pS is known from Pantoea bacteria (WO98 / 59054), plasmids capable of improving microbial growth under acidic conditions when transformed into microorganisms are unknown

Method used

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  • Novel plasmid autonomously replicable in enterobacteriaceae family
  • Novel plasmid autonomously replicable in enterobacteriaceae family
  • Novel plasmid autonomously replicable in enterobacteriaceae family

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Embodiment 1

[0120] Because pEA320 is very large (approximately 320 kbs), it is difficult to isolate as circular DNA and difficult to use to transform strains. In addition, since neither a drug resistance gene nor a marker capable of dominant selection exists on the plasmid, it is necessary to introduce a marker gene to make dominant selection possible. Therefore, pUT399 was constructed to enable homologous recombination to occur, and thus, the strain could be selected using chloramphenicol (US Patent Application Publication No. 20040180404).

[0121] pUT399 was constructed as follows. The origin of replication of R6K and the mob of RP4 were amplified by PCR using pUT-miniTn5-Cm (which can be purchased from Biomedal) as template and P7 (SEQ ID NO: 9) and P8 (SEQ ID NO: 10) as primers Gene. In addition, when pHSG399 (TaKaRa) was used as template, and P9 (SEQ ID NO: 11) and P10 (SEQ ID NO: 12) were used as primers, a gene containing chloramphenicol resistance and a multiple cloning site we...

Embodiment 2

[0126]

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Abstract

The present invention describes a gene isolated from a microorganism selected from bacteria of the genera Pantoea, bacteria of the genera Erwinia, and bacteria of the genera Enterobacter, and encoding a Rep protein or a homologue thereof. The invention also describes a plasmid containing this gene, which is autonomously replicable in Enterobacteriaceae. The invention also describes Enterobacteriaceae microorganisms containing the plasmid.

Description

technical field [0001] The present invention relates to novel plasmids, which are autonomously replicable in microorganisms from the family Enterobacteriaceae, and to methods of using the plasmids and microorganisms containing the plasmids. Background technique [0002] Currently, the cultivation and improvement of microorganisms from the family Enterobacteriaceae, including Escherichia coli, by DNA recombinant technology is underway. In order to breed and improve microorganisms by DNA recombination technology, plasmids and broad host spectrum vectors derived from microorganisms of different genera are often used. Generally, however, plasmids from microorganisms with similar properties are used. [0003] Plasmids derived from microorganisms of the family Enterobacteriaceae include small plasmids such as pBR322, RSF1010 (Gene, vol.75, (2) pp.271-288, 1989) and pUC (Gene, Oct. 1982, 1989) all derived from Escherichia coli. (3): 259-68), pHCM2 derived from Salmonella (Salmone...

Claims

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Application Information

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IPC IPC(8): C07K14/27C12N15/63
CPCC12P13/04C07K14/24C07K14/27C12N15/63
Inventor 原吉彦泉井裕野口博美
Owner AJINOMOTO CO INC
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