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Primer composition for detecting or assisting in detecting chlamydia psittaci and application of primer composition

A kind of technology of Chlamydia psittaci, primer composition, applied in the primer composition of detection or auxiliary detection of Chlamydia psittaci and its application field, can solve the problem that can not be adapted to be simple, fast, sensitive, the sensitivity of detection needs to be improved, and the reaction time is long. and other problems to achieve the effect of high sensitivity, high specificity and strong specificity

Active Publication Date: 2022-05-27
ACADEMY OF MILITARY MEDICAL SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0003] Serological diagnosis is mainly based on indirect immunofluorescence, but this method can only detect Chlamydia psittaci antibodies 1-2 weeks after the onset of the disease, and the timeliness is poor
Gene detection technology based on the principle of PCR is the mainstream method for the molecular biology diagnosis of Chlamydia psittaci, but the detection requires the use of professional instruments, the reaction time is also long, and the sensitivity of detection needs to be improved, and it cannot adapt to the clinical "simple and fast" method. , Sensitive” requirements

Method used

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  • Primer composition for detecting or assisting in detecting chlamydia psittaci and application of primer composition
  • Primer composition for detecting or assisting in detecting chlamydia psittaci and application of primer composition
  • Primer composition for detecting or assisting in detecting chlamydia psittaci and application of primer composition

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0064] Example 1. Design and screening of Chlamydia psittaci primers and probes.

[0065] (1) Design of primers and probes.

[0066] Through file retrieval, the inventors analyzed and determined that the specific sequence in the CPSIT_RS02830 gene of Chlamydia psittaci in the present invention is the target gene. The known template gene sequence, that is, the nucleotide sequence shown in SEQ ID NO: 1, was obtained from the NCBI database. According to the RAA primer and probe design principle, 6 primers and 2 probes were designed. The sequences are as follows Figure 8 shown. The probe consists of a fluorescein (such as carboxyfluorescein FAM), a 5'-end sequence, tetrahydrofuran (THF), a 3'-end sequence, and a 3'-end extension blocking group (such as a phosphate group). The length of the probe is 45-60 bp, of which the 5' end is at least 30 bp and the 3' end is at least 15 bp.

[0067] The probe of the present invention is labeled with fluorescein FAM at the 5' end, and an ...

Embodiment 2

[0080] Example 2: Sensitivity evaluation of RAA detection.

[0081] Dilute Chlamydia psittaci genomic DNA to 10 5 to 10 0 A series of different concentrations, such as 1 copy / μL, were added to the reaction system determined in Example 1 (that is, the reaction temperature was 39 °C and the amplification time was 15 min), and the selected primer probe set (the primer probe No. 12) was used. Group), using the amplification and detection conditions determined in Example 1 to perform RAA detection on the above templates with different copy numbers to observe the sensitivity of RAA detection. A set of negative controls were set up. The negative controls did not add template, and the template volume was supplemented with water.

[0082] As a result, results such as image 3 shown, image 3 The template concentrations in lanes numbered 1-6 were 1 × 10 5 copies / μL, 1×10 4 copies / μL, 1×10 3 copies / μL, 1×10 2 copies / μL, 1×10 1 copies / μL, 1×10 0 copies / μL, 7 represents NC negati...

Embodiment 3

[0083] Example 3: Specificity evaluation of RAA detection.

[0084] Specificity was evaluated by Anaplasma phagocytogenes, Rickettsia rickettsiae, Rickettsia Heilongjiang, Rickettsia Siberia, Rickettsia Canada, Rickettsia australis, Rickettsia Morsei, Rickettsia Platt. Rickettsia, Henzerling strain of C. bainii, seven strains of C. bainii, Legionella pneumophila, Ehrlichia chaffee, Listeria monocytogenes, Salmonella typhimurium, aureus The genomic DNAs of Staphylococcus, Vibrio cholerae, Bartonella 5 days, Salmonella typhi, Streptococcus suis, Shigella sonnei and Chlamydia trachomatis were used as controls to determine the specificity of the RAA detection method of the present invention.

[0085] Anaplasma phagocytosis, Rickettsia rickettsii, Rickettsia Heilongjiang, Rickettsia Siberia, Rickettsia Canada, Rickettsia australis, Rickettsia Przewalskii, Rickettsia Przewalskii Chlorella, Henzerling strain of C. bainii, seven strains of C. bainii, Legionella pneumophila, Ehrlichie...

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Abstract

The invention discloses a composition for detecting or assisting in detecting chlamydia psittaci and application of the composition, and belongs to the field of compositions used in a detection method containing nucleic acid or microorganisms. The technical problem to be solved by the invention is how to detect or assist in detecting chlamydia psittaci with high sensitivity and / or high specificity and / or quickly / simply. The invention provides a primer composition for detecting or assisting in detecting chlamydia psittaci. The primer composition is composed of single-stranded DNA molecules with nucleotide sequences as shown in sequences 2 and 3 in a sequence table. When the composition for detecting or assisting in detecting chlamydia psittaci is used for detecting the chlamydia psittaci through RAA amplification, rapidness and high efficiency are achieved, the amplification reaction can be completed within 15 minutes, and a large number of amplification reactions can be completed only at the constant temperature; the detection result is good in specificity and high in sensitivity; the identification method is simple and convenient, and is suitable for rapid detection of on-site or clinical specimens.

Description

technical field [0001] The present invention relates to a primer composition for detecting or assisting the detection of Chlamydia psittaci and its application in the field of compositions used in the detection method comprising nucleic acid or microorganism. Background technique [0002] Chlamydia psittaci ( Chlamydia psittaci ) belongs to the order Chlamydia ( Chlamydiales ), Chlamydia ( Chlamydiaceae ), Chlamydia ( Chlamydia ), a Gram-negative, strictly intracellular parasitic microorganism, the size is between bacteria and viruses, mainly transmitted by air and contact, and is a kind of biological warfare agent. Parrots were initially thought to be the host of the pathogen, and the disease caused by it was called psittacosis, and humans, birds and some mammals are susceptible. Birds or poultry with bacteria, bacteria-containing secretions or excreta contaminated environment, feathers and dust can be the source of infection. Human infection with psittacosis is usu...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/689C12Q1/6844C12N15/11C12R1/01
CPCC12Q1/689C12Q1/6844C12Q2521/507C12Q2522/101Y02A50/30
Inventor 焦俊欧阳譞熊小路何培圣于永慧付梦姣赵明亮
Owner ACADEMY OF MILITARY MEDICAL SCI
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