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Oligonucleotide aptamer group for specifically identifying staphylococcus aureus and use thereof

A staphylococcus and oligonucleotide technology, applied in the application field of identifying staphylococcus aureus, can solve difficult and time-consuming problems, achieve the effects of low preparation cost, simple operation, and improved sensitivity and specificity

Active Publication Date: 2010-03-10
中国人民解放军军事医学研究院
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Problems solved by technology

Finding these differences and producing specific antibodies is obviously a time-consuming and difficult task

Method used

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  • Oligonucleotide aptamer group for specifically identifying staphylococcus aureus and use thereof
  • Oligonucleotide aptamer group for specifically identifying staphylococcus aureus and use thereof
  • Oligonucleotide aptamer group for specifically identifying staphylococcus aureus and use thereof

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Embodiment Construction

[0016] The present invention will be further described below by subtractive SELEX screening to obtain Staphylococcus aureus-specific oligonucleotide aptamers and comparing the recognition and detection rates of five specific oligonucleotide aptamers to Staphylococcus aureus.

[0017] 1. Synthesize a random single-stranded DNA library and primers (completed by Invitrogen).

[0018] Library GP45: 5'-GCAATGGTACGGTACTTCC(45N)CAAAAGTGCACGCTACTTTGCTAA-3'

[0019] 5' Primer (Plong-1): 5'-GCAATGGTACGGTACTTCC-3'

[0020] 3' Primer (P11): 5'-TTAGCAAAGTAGCGTGCACTTTTG-3'

[0021] 3' Primer (Pstem-loop3): 5'-GCTAAGCGGGTGGGACTTCCTAGTCCCACCCGCTTAGCAAAGTAGCGTGCACTTTTG-3'

[0022] 2. Acquisition and processing of bacterial targets for screening.

[0023] Staphylococcus aureus, Streptococcus and Staphylococcus epidermidis were cultured in TSB medium, and they were shaken at 37°C until early logarithmic growth (OD 600 about 0.2). Then collect the bacteria separately, take 100μl bacterial so...

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Abstract

The invention belongs to the technical field of biotechnology and relates to the sequences of five oligonucleotide aptamers for specifically identifying staphylococcus aurei and use thereof. The fivesinge-chain DNA oligonucleotide aptamers for specifically identifying the staphylococcus aureus are obtained by a bacterial subtraction SELEX technology. After FITC is marked, the five aptamers are proved by fluorescent staining to be capable of specifically identifying the strain of the staphylococcus aurei. After being used in combination, the five ligands can improve sensibility in identification of target bacteria and the ratios of detected different staphylococcus aureus strains and the detected staphylococcus aurei in different growth states. Therefore, the sequences of the five aptamershave wide application prospects in quick detection of the staphylococcus aurei in samples including suspected terrorist attack biological agents and identification of the staphylococcus aurei and other gram-positive cocci.

Description

Field of invention: [0001] The invention relates to the acquisition of a group of oligonucleotide aptamers specifically recognizing staphylococcus aureus and the application of these oligonucleotide aptamers in identifying staphylococcus aureus. Background technique: [0002] Bacteria are common pathogenic factors of clinical diseases and seriously threaten human health. How to quickly and sensitively identify the types and properties of important pathogenic microorganisms is a major topic that countries all over the world have invested a lot of manpower and material resources to study. The traditional method of detecting pathogenic bacteria often needs to isolate pathogenic microorganisms first, then culture them, and then identify them by classical methods. Time-consuming and insensitive are common problems of these methods. It is necessary to develop rapid and sensitive detection of pathogenic microorganisms, especially in the prevention of bioterrorism attacks. Althou...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/68C12Q1/14C12N15/11
Inventor 邵宁生李少华曹晓晓丁红梅黄艳萍徐华高亚萍王芳杨光刘农乐赵强
Owner 中国人民解放军军事医学研究院
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