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Probe, method and kit used for absolute quantification of bacterial microorganisms, and application of kit

A technology of microorganisms and detection kits, applied in biochemical equipment and methods, measurement/inspection of microorganisms, DNA/RNA fragments, etc., which can solve the problems of unsuitable and timely detection, insufficient detection accuracy, and the inability to realize the detection of total bacterial microorganisms And other issues

Pending Publication Date: 2021-03-26
JIANGNAN UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0006] A method, kit and application for the absolute quantification of bacterial microorganisms of the present invention solve at least one of the following technical problems: (1) the existing method cannot realize the total detection of all bacterial microorganisms; (2) the existing Quantitative methods have low species resolution and / or insufficient detection accuracy; (3) Existing quantitative methods require high-cost instruments and / or strict operating environments, and are not suitable for timely detection after production sampling; (4) Existing quantitative methods are cumbersome to operate, etc.

Method used

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  • Probe, method and kit used for absolute quantification of bacterial microorganisms, and application of kit
  • Probe, method and kit used for absolute quantification of bacterial microorganisms, and application of kit
  • Probe, method and kit used for absolute quantification of bacterial microorganisms, and application of kit

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0061] Example 1: Bacterial microorganism quantitative probe combination reagent

[0062] Probe combination reagent; contains independently packaged signal probe reagent and quenching probe reagent; wherein, the signal probe sequence is shown in SEQ ID NO.1 or SEQ ID NO.3, and the quenching probe sequence is SEQ ID NO.2 (corresponding to the signal probe SEQ ID NO.1) or SEQ ID NO.4 (corresponding to the signal probe SEQ ID NO.3).

[0063] Signal probe reagents and quench probe reagents, in dry powder or liquid form; in dry powder form, they can be diluted to an appropriate concentration before the experiment, for example, use sterile water or buffer to dilute to a concentration of 20 μM; in liquid form , the concentration can be 20-200μM, the reagent can be diluted before use, or used directly.

Embodiment 2

[0064] Example 2: Bacterial microorganism quantitative kit and its use

[0065] Bacterial microorganism quantification kit, containing independently packaged signal probe reagent and quenching probe reagent; wherein, the signal probe sequence is shown in SEQ ID NO.1 or SEQ ID NO.3, and the quenching probe sequence is shown in SEQ ID NO. It is shown in SEQ ID NO.2 (corresponding to signal probe SEQ ID NO.1) or SEQ ID NO.4 (corresponding to signal probe SEQ ID NO.3).

[0066] When the kit is used, it can be mixed with heme, buffer, 2,2-azino-bis-(3-ethylbenzodihydrothiazoline-6-sulfonic acid) diammonium salt (ABTS), H 2 O 2 With the use of.

[0067] The usage method is:

[0068] (1) Solution configuration. Prepare 100nM heme solution (reagent 1); prepare Tris-HCL with a final concentration of 50mM, KCl with a final concentration of 50mM, and a final pH of 7.9 (reagent 2); 7mM of 2,2-azino-bis-( 3-ethylbenzodihydrothiazoline-6-sulfonic acid) diammonium salt (ABTS) (reagent 3...

Embodiment 3

[0075] Example 3: Bacterial microorganism quantitative kit

[0076] Bacterial microorganism quantification kit, containing independently packaged signal probe reagent and quenching probe reagent; wherein, the signal probe sequence is shown in SEQ ID NO.1 or SEQ ID NO.3, and the quenching probe sequence is shown in SEQ ID NO. It is shown in SEQ ID NO.2 (corresponding to signal probe SEQ ID NO.1) or SEQ ID NO.4 (corresponding to signal probe SEQ ID NO.3).

[0077] The kit also contains 100 nM heme solution (reagent 1), Tris-HCL buffer, 7 mM 2,2-azino-bis-(3-ethylbenzodihydrothiazoline-6-sulfonic acid) ) diammonium salt (ABTS), 7mM H 2 O 2 solution.

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Abstract

The invention discloses a probe, a method and a kit used for absolute quantification of bacterial microorganisms, and application of the kit, and belongs to the biological field, the fermentation field and the detection field. The probe and the kit used for the quantification of the bacterial microorganisms can realize the total quantity detection of all bacterial microorganisms, do not need to use expensive instruments during detection and the quantification of the bacterial microorganisms, and can quickly finish quantification work in 2.5h. Meanwhile, a sample used by the invention does notneed to carry out nucleic acid extraction. When the probe and the detection kit disclosed by the invention are used for the quantification of the bacterial microorganisms, the probe and the detectionkit have the characteristics of being quick, convenient, cheap and accurate.

Description

technical field [0001] The invention relates to a probe, method, kit and application for absolute quantification of bacterial microorganisms, belonging to the fields of biology, fermentation and detection. Background technique [0002] There are complex microbial flora involved in the brewing process of traditional fermented food, among which bacteria are important functional microorganisms. These microorganisms are closely related to the quality of the final product. During the fermentation process, bacterial microorganisms participate in a series of physiological and biochemical reactions related to substrate degradation and product synthesis, converting macromolecules such as available glycogen and protein into alcohols. , aldehydes, acids, esters and other flavor components, giving products unique flavor and sensory characteristics. Therefore, real-time tracking of the growth trend of bacterial microorganisms during the fermentation process has important guiding signifi...

Claims

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Application Information

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IPC IPC(8): C12Q1/689C12Q1/6816C12Q1/06C12N15/11
CPCC12Q1/689C12Q1/6816C12Q2521/50C12Q2525/301Y02A50/30
Inventor 吴群徐岩杜如冰
Owner JIANGNAN UNIV
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