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CN-122012671-A - Tongue swab sample preservation solution and application thereof

CN122012671ACN 122012671 ACN122012671 ACN 122012671ACN-122012671-A

Abstract

The application belongs to the technical field of nucleic acid detection, and particularly relates to a tongue swab sample preservation solution and application thereof. Comprising 20mM-100mM Tris-HCl or HEPES, and 0.3% -0.6% (v/v) Tween 20, 0.04% -0.08% (v/v) Triton X-100, 0.03% -0.05% (w/v) guanidine isothiocyanate, 0.1mM-0.3mM EDTA-2Na, 0.1wt% -0.2wt% sucrose and 20mM-25mM NaOH. The pH value of the tongue swab sample preservation solution is 7-8, and the tongue swab sample preservation solution can be adapted to enrichment magnetic beads, so that effective extraction of low-abundance mycobacterium tuberculosis nucleic acid in the tongue swab sample is realized.

Inventors

  • CHEN JIAMIN
  • HUANG SHAOYA
  • WANG JINHUI
  • CHEN XINYU
  • HE LI
  • DING FENG
  • WU KANG
  • DAI LIZHONG

Assignees

  • 圣湘生物科技股份有限公司

Dates

Publication Date
20260512
Application Date
20251011

Claims (10)

  1. 1. A tongue swab sample preservation solution comprising 20mM-100mM Tris-HCl or HEPES, and 0.3% -0.6% (v/v) tween 20, 0.04% -0.08% (v/v) Triton X-100, 0.03% -0.05% (w/v) guanidine isothiocyanate, 0.1mM-0.3mM EDTA-2Na, 0.1% -0.2% sucrose and 20mM-25mM NaOH.
  2. 2. The tongue swab sample preservation solution according to claim 1, comprising 45mM-55mM Tris-HCl, 0.3% -0.4% (v/v) tween 20, 0.04% -0.06% (v/v) Triton X-100, 0.03% -0.05% (w/v) guanidine isothiocyanate, 0.1mM-0.2mM EDTA-2Na, 0.1% -0.2wt% sucrose and 20mM-25mM NaOH.
  3. 3. A nucleic acid extraction product of mycobacterium tuberculosis, comprising: the tongue swab sample preservation solution according to any one of claims 1 to 2, or One or more of magnetic beads and swabs, and the tongue swab sample preservation solution.
  4. 4. A nucleic acid extraction product of mycobacterium tuberculosis according to claim 3, wherein the nucleic acid extraction product satisfies one or more of the following conditions: (1) The magnetic beads comprise one or more of a silicon hydroxyl modified magnetic bead and an amino modified magnetic bead, optionally the magnetic beads comprise an amino modified magnetic bead, and (2) The swab comprises one or more of a sponge swab, a flocked swab, and a different breakpoint swab, optionally the swab comprises a different breakpoint swab.
  5. 5. A kit for detecting mycobacterium tuberculosis, the kit is characterized by comprising: 1) The tongue swab sample preservation solution according to any one of claims 1 to 2 or the nucleic acid extraction product according to any one of claims 3 to 4, and 2) A nucleic acid detection reagent; optionally, the nucleic acid detection reagent is a lyophilized reagent.
  6. 6. The detection kit for mycobacterium tuberculosis according to claim 5, wherein the nucleic acid detection reagent comprises a PCR detection reagent; optionally, the nucleic acid detection reagent comprises one or more of primers, probes, PCR buffers, dNTPs, a hot start enzyme, and a UDG enzyme.
  7. 7. The kit for detecting mycobacterium tuberculosis according to claim 6, wherein the nucleic acid detection reagent detects IS6110 gene and/or IS1081 gene; Optionally, the nucleic acid detection reagent comprises one or more of the following primer probes: primer probes shown in SEQ ID No.2 to SEQ ID No.4, Primer probes shown in SEQ ID No.5 to SEQ ID No.7, Primer probes shown in SEQ ID No.8 to SEQ ID No.10, Primer probes shown in SEQ ID No.12 to SEQ ID No.14, Primer probes shown in SEQ ID No.15 to SEQ ID No.17, Primer probes shown in SEQ ID No.18 to SEQ ID No.20, and Primer probes shown in SEQ ID No.22 to SEQ ID No. 24.
  8. 8. A method of processing a mycobacterium tuberculosis lingual swab sample, the method comprising: placing a tongue swab sample to be tested in the tongue swab sample preservation solution according to any one of claims 1 to 2, and preparing a sample liquid to be tested.
  9. 9. The method for treating a mycobacterium tuberculosis tongue swab sample according to claim 8, wherein the method employs the nucleic acid extraction product of any one of claims 3 to 4; optionally, the treatment method further comprises the steps of carrying out ultrasonic treatment on the sample liquid to be detected, adding the magnetic beads for nucleic acid enrichment, collecting the magnetic beads for adsorbing nucleic acid, and preparing nucleic acid to be detected; optionally, the ultrasonic conditions comprise frequency of 35-38kHz, power of 80% -90%, pressure of 170kPa-190kPa, or/and time of 1.5-3min; Alternatively, 1-2 mu L of the magnetic bead suspension with the concentration of 2.5-5 mg/mL is added into 1mL of the sample liquid to be tested.
  10. 10. A method for detecting a mycobacterium tuberculosis lingual swab sample, the method comprising: treating a tongue swab sample to be tested with the treatment method according to any one of claims 8 to 9, and detecting; Optionally, the detection is performed using a nucleic acid detection reagent as defined in any one of claims 5 to 7.

Description

Tongue swab sample preservation solution and application thereof Technical Field The application belongs to the technical field of nucleic acid detection, and particularly relates to a tongue swab sample preservation solution and application thereof. Background Mycobacterium tuberculosis complex (Mycobacterium tuberculosis complex, TBC) is a group of pathogens capable of causing tuberculosis, mainly including Mycobacterium tuberculosis, mycobacterium bovis, mycobacterium africanum, mycobacterium tenuifolia, etc. Among them, mycobacterium tuberculosis, mycobacterium bovis and Mycobacterium africanus are all pathogenic to human, and it is most common to form tuberculosis by invasion of the whole body organs through respiratory tract, digestive tract and damaged skin mucosa and pulmonary involvement. In tuberculosis prevention and control work, a rapid and accurate diagnosis technology is particularly important. The techniques not only can accelerate the treatment process of patients, reduce the pain caused by diseases, but also can effectively inhibit the spread of the diseases and protect more people from being damaged. At present, xpert MTB/RIF Ultra sputum detection occupies an important position in the tuberculosis diagnosis field with excellent sensitivity and specificity. However, the limitations of this approach are not negligible-it requires the patient to provide a sputum sample, a condition that constitutes a significant challenge for children, HIV co-infected persons, and special populations of critically ill patients. Therefore, the search and development of novel, more convenient and easy to operate diagnostic techniques is particularly urgent, wherein molecular nucleic acid detection is a better diagnostic protocol due to timeliness, high specificity and sensitivity. In recent years, non-invasive tongue swab sample PCR nucleic acid detection methods have received attention. The method greatly simplifies the sample collection process, provides more comfortable detection experience for patients, and is particularly suitable for patients who cannot provide sputum samples. Preliminary researches show that tongue swab PCR detection is equivalent to the existing sputum reference test in terms of sensitivity and specificity, and shows great application potential in tuberculosis diagnosis. The real-time fluorescence PCR technology is characterized in that a specific primer probe is designed to dynamically monitor the change of a fluorescence signal and automatically analyze the result in real time, so that the method is a specific, time-saving and convenient detection means and can accurately and efficiently detect the mycobacterium tuberculosis. The limitation of the current qPCR technology for tuberculosis tongue swabs is mainly that the number of pathogens in tongue swabs is small, the abundance is insufficient, and the detection is difficult. In view of this, the present application has been made. Disclosure of Invention In view of this, one or more embodiments of the present application provide a tongue swab sample preservation solution and its use. The method comprises the following technical scheme: One or more embodiments of the present application provide a tongue swab sample preservation solution comprising 20mM-100mM Tris-HCl or HEPES, and 0.3% -0.6% (v/v) Tween 20, 0.04% -0.08% (v/v) Triton X-100, 0.03% -0.05% (w/v) guanidine isothiocyanate, 0.1mM-0.3mM EDTA-2Na, 0.1wt% -0.2wt% sucrose, and 20mM-25mM NaOH. In some embodiments of the application, the tongue swab sample preservation solution comprises 45mM-55mM Tris-HCl,0.3% -0.4% (v/v) Tween 20, 0.04% -0.06% (v/v) Triton X-100, 0.03% -0.05% (w/v) guanidine isothiocyanate, 0.1mM-0.2mM EDTA-2Na, 0.1% -0.2% sucrose, and 20mM-25mM NaOH. One or more embodiments of the present application provide a nucleic acid extraction product of mycobacterium tuberculosis, comprising the tongue swab sample preservation solution, or one or more of a magnetic bead and a swab, and the tongue swab sample preservation solution. In some embodiments of the application, the nucleic acid extraction product satisfies one or more of the following conditions: (1) The magnetic beads comprise one or more of a silicon hydroxyl modified magnetic bead and an amino modified magnetic bead, optionally the magnetic beads comprise an amino modified magnetic bead, and (2) The swab comprises one or more of a sponge swab, a flocked swab, and a different breakpoint swab, optionally the swab comprises a different breakpoint swab. One or more embodiments of the present application provide a kit for detecting mycobacterium tuberculosis, which comprises 1) the tongue swab sample preservation solution or the nucleic acid extraction product, and 2) a nucleic acid detection reagent, and optionally, the nucleic acid detection reagent is a freeze-dried reagent. In some embodiments of the application, the nucleic acid detection reagent comprises a PCR detection reagent, optionally the nucleic aci