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Nucleic acid identification sequence and detection method for singular proteus

A technology of Proteus mirabilis and identification sequence, which is applied in the field of nucleic acid identification sequence and detection of Proteus mirabilis, can solve the problem of no specific nucleic acid sequence, etc., and achieves the effect of high conservation and specificity and broad application prospects.

Inactive Publication Date: 2010-12-22
INST OF PLA FOR DISEASE CONTROL & PREVENTION
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

So far, there is little sequence information about Proteus mirabilis, and there is no specific nucleic acid sequence that can be used for identification

Method used

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  • Nucleic acid identification sequence and detection method for singular proteus
  • Nucleic acid identification sequence and detection method for singular proteus

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Experimental program
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Effect test

Embodiment 1

[0039] Example 1, Screening of Proteus mirabilis specific nucleic acid marker sequence

[0040] The following method is used to screen the specific nucleic acid identification sequence of Proteus mirabilis, and the specific process includes the following steps:

[0041] 1. Construction of Proteus mirabilis genome library

[0042] 1. Extraction of Proteus mirabilis genomic DNA

[0043] Proteus mirabilis preserved in glycerol (strain number: 82009, purchased from China National Institute for the Control of Pharmaceutical and Biological Products) was streaked and inoculated on LB plates, cultured at 37°C for 24 hours, and single colonies were picked and inoculated on In 5mL LB liquid medium, the bacteria grow to the mid-logarithmic growth phase (OD 600 Value is 0.8), centrifuged to collect the thalli, use the genome extraction kit (purchased from Promega Company) and refer to the kit manual to extract the genomic DNA of the thalline, then dissolve the extracted genomic DNA with...

Embodiment 2

[0064] Embodiment 2, the common PCR detection of Proteus mirabilis

[0065] Common PCR detection primer PRM0007-F (upstream primer) designed according to the specific nucleic acid identification sequence PRM0007 of Proteus mirabilis obtained in Example 1: 5'-ACGGAATGAAAGGAGTC-3' (sequence 2 in the sequence listing) and PRM0007-R (downstream Primer): 5'-CACCAGCAAGAATAGAACA-3' (sequence 3 in the sequence listing) to the standard strain of Proteus mirabilis (Proteus mirabilis 82009, purchased from China Institute for the Control of Pharmaceutical and Biological Products) and other bacterial strains (Wang Yong et al., Journal of Preventive Medicine of the People's Liberation Army , 2007, 25 (2): 94-97) for PCR detection, the specific detection method is as follows:

[0066] Using the genomic DNA of each strain to be tested as a template, PCR amplification was carried out under the guidance of primers PRM0007-F and PRM0007-R, and the PCR reaction system (25 μl) was: 1×PCR buffer (i...

Embodiment 3

[0067] Embodiment 3, the SYBR Green I real-time fluorescent quantitative PCR detection of Proteus mirabilis

[0068] Design SYBR GreenI real-time fluorescent quantitative PCR detection primer PRM0007-F1 (upstream primer) according to the specific nucleic acid identification sequence PRM0007 of Proteus mirabilis obtained in Example 1: 5'-ATGAATACCGTGCCCAAGA-3' (sequence 4 in the sequence listing) and PRM0007-R1 (Downstream primer): 5'-CACCAGCAAGAATAGAACAAGA-3' (sequence 5 in the sequence table) to be tested (Proteus mirabilis, strain number 82009, purchased from China Institute for the Control of Pharmaceutical and Biological Products) and other bacterial strains (Wang Yong et al., People's Liberation Army Prevention Medical Journal, 2007, 25 (2): 94-97) carries out SYBR Green I quantitative PCR detection, and specific detection method is as follows:

[0069] Using the genomic DNA of the strain to be tested as a template, under the guidance of primers PRM0007-F and PRM0007-R, P...

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Abstract

The invention discloses a specific nucleic acid marking sequence of proteus mirabilis, a PCR detecting method and the application. The aim of the invention is to provide the specific nucleic acid marking sequence of the proteus mirabilis, a qualitative and quantitative PCR detecting method of the proteus mirabilis and the application in preparing qualitative and quantitative PCR detecting kits ofthe proteus mirabilis. The specific nucleic acid marking sequence of the proteus mirabilis is one of following nucleic acid sequences: firstly, a DNA sequence of SEQ ID NO: 1 in the table, secondly, a restricted DNA sequence of the SEQ ID NO: 1 in the table, which has the homology more than 90% and is the specific nucleic acid sequence of the proteus mirabilis, and thirdly, a nucleic acid sequence which can cross-breed with the restricted DNA sequence of the SEQ ID NO: 1 in the table under the high-stringent condition. The invention can be used in qualitative and quantitative molecular detection of the proteus mirabilis (including the detecting methods of ordinary PCR, quantitative PCR, chips, hybrid and the like) and has broad application prospect.

Description

technical field [0001] The present invention relates to the specific nucleic acid identification sequence of Proteus mirabilis and its detection method and application, in particular to the specific nucleic acid identification sequence of Proteus mirabilis and the qualitative and quantitative PCR detection of Proteus mirabilis and the qualitative and quantitative PCR detection of Proteus mirabilis in preparation application in the kit. Background technique [0002] Proteus species exists in water, soil, decayed organic matter, and the intestinal tract of humans and animals. It is an opportunistic pathogen and often causes secondary infections, such as chronic otitis media, trauma infections, etc. Diseases such as diarrhea and food poisoning (Coker C, Poore CA, Li X et al. Pathogenesis of Proteus mirabilis urinary tract infection[J]. Microbes Infect, 2000, 2: 1497-1505.; Mobley HL, and Belas R. Swarming and pathogenicity of Proteusmirabilis in the urinary tract [J]. Trends M...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N15/31C12Q1/68C12Q1/04C07H21/04
Inventor 陈泽良赵瑾王玉飞杜昕颖乔凤张玲王勇张伶黄留玉
Owner INST OF PLA FOR DISEASE CONTROL & PREVENTION
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