Primer, probe, kit and detection method for multiple detection of chlamydia trachomatis, neisseria gonorrhoeae and ureaplasma

A technology for multiple detection of Chlamydia trachomatis, applied in biochemical equipment and methods, recombinant DNA technology, and microbial determination/inspection, etc., can solve the problems of increasing detection cost and complicated operation, difficult to meet early diagnosis, and high technical requirements, Achieve the effect of improving the tedious operation, reducing the high detection cost, and ensuring high efficiency

Inactive Publication Date: 2019-05-31
中生方政生物技术股份有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0007] 1) The isolation and culture of pathogens is the gold standard for CT / NG / UU, but the isolation and culture method requires high technical requirements, is expensive, and takes a long time, so it is rarely used in clinical testing
[0008] 2) There is still lag in immunological diagnosis, which cannot accurately reflect whether the current infection or pathogen is carried, and it is difficult to meet the requirements of early diagnosis
[0011] At present, most of the products adopt a single fluorescent quantitative PCR detection method for CT / NG / UU. It is necessary to conduct multiple detections on a clinical sample before drawing conclusions separately, which increases the detection cost and cumbersome operation.

Method used

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  • Primer, probe, kit and detection method for multiple detection of chlamydia trachomatis, neisseria gonorrhoeae and ureaplasma
  • Primer, probe, kit and detection method for multiple detection of chlamydia trachomatis, neisseria gonorrhoeae and ureaplasma
  • Primer, probe, kit and detection method for multiple detection of chlamydia trachomatis, neisseria gonorrhoeae and ureaplasma

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0067] Embodiment 1: be used for the design of the primer probe pair of rapid detection CT / NG / UU

[0068] Download the sequences of Chlamydia trachomatis cryptic plasmamid, Neisseria gonorrhoeae plasmamid, Ureaplasma 16SrRNA, and HBB gene sequences in NCBI, and design primer pairs and probes. The sequences are as follows:

[0069] Table 1 Primer and Probe Sequences

[0070]

[0071]

Embodiment 2

[0072] Embodiment 2: The establishment of the real-time fluorescent quantitative PCR kit for rapid detection CT / NG / UU

[0073] Real-time fluorescent quantitative PCR kit for rapid detection of human CT / NG / UU, including reaction mixture, sample extract, positive control, negative control, instructions and box.

[0074] The reaction mixture contains upstream and downstream primers and probes (SEQ ID NO: 1-12), Anstart qPCR Master Mix enzyme mixture, Anstart qPCR Master Mix 5× reaction buffer.

[0075]Among them, Anstart qPCR Master Mix Enzyme Mixture and Anstart qPCR Master Mix5×Reaction Buffer are provided by Feipeng Biological Co., Ltd. Anstart qPCR Master Mix Enzyme Mixture is diluted 25 times for use, and Anstart qPCR Master Mix5×Reaction Buffer is diluted 5 times for use .

[0076] The primers CT-F, CT-R, Ng-F, Ng-R, UU-F, UU-R, HBB-F, and HBB-R have a final concentration of 500 nM.

[0077] The concentration of probes CT-P, Ng-P, UU-P and HBB-R is 200 nM.

[0078] Among...

Embodiment 3

[0090] Embodiment 3: the rapid detection method of CT / NG / UU nucleic acid detection kit

[0091] Utilize the kit of embodiment 2 to rapidly detect CT / NG / UU in human vaginal secretions and urinary tract secretions, the specific steps are as follows:

[0092] (1) Nucleic acid extraction: samples of vaginal secretions (5 copies, No. 1-5), urinary tract secretions (No. 6-10) (use the culture method to determine whether they are infected by CT, NG or UU, which is the current industry standard Gold standard) Add 1 mL of normal saline to the collection tube, shake and wash the cotton swab fully, then squeeze the cotton swab against the wall and discard, transfer 500 μL of the liquid to a 1.5 mL centrifuge tube, centrifuge at 13,000 rpm for 5 minutes, discard the supernatant, Add 1 mL of normal saline to the precipitate, break up the precipitate, centrifuge at 13,000 rpm for 5 minutes, discard the supernatant, add 50 μL of the sample extract solution that has been shaken and mixed to t...

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Abstract

The invention relates to the technical field of detection of pathogenic microorganisms, in particular to primers, a probe, a kit and a detection method for multiple detection of chlamydia trachomatis,neisseria gonorrhoeae and ureaplasma. The kit comprises a CT / NG / UU conserved region-specific primer pair and the Taqman fluorescent probe. The kit can accurately detect CT / NG / UU infected samples by real-time fluorescent quantitative PCR, and is convenient and quick to use, high in sensitivity, good in specificity and high in repeatability.

Description

technical field [0001] The invention relates to the technical field of detection of pathogenic microorganisms, in particular to primers, probes, kits and detection methods for multiple detection of Chlamydia trachomatis, Neisseria gonorrhoeae and Ureaplasma. Background technique [0002] Sexually transmitted diseases (STD) refer to a group of infectious diseases that can be transmitted through sexual contact. In China, people call it STD for short. As a social disease, STD not only causes different degrees of physical damage to patients, but also brings mental pain, and also brings misfortune to spouses, children, and families, causing serious losses to society and the country. In patients infected with STD diseases, males can cause epididymitis, spermatic cord inflammation and infertility, and females can cause pelvic inflammatory disease, salpingitis, endometritis, ectopic endometritis, ectopic pregnancy, and miscarriage. Among them, Chlamydia trachomatis (CT), Neisseria ...

Claims

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Application Information

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IPC IPC(8): C12Q1/689C12Q1/6851C12Q1/04C12N15/11
Inventor 蔺皓邹国宝魏颖颖宋高尚吴茜刘欣欣沈江卫
Owner 中生方政生物技术股份有限公司
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