CN116949161A - Group of tuberculosis serum exosome miRNA markers and application thereof - Google Patents
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- 201000008827 tuberculosis Diseases 0.000 title claims abstract description 57
- 210000001808 exosome Anatomy 0.000 title claims abstract description 23
- 210000002966 serum Anatomy 0.000 title claims abstract description 23
- 108091070501 miRNA Proteins 0.000 title claims description 5
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- 238000002123 RNA extraction Methods 0.000 description 3
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- 208000035473 Communicable disease Diseases 0.000 description 1
- 241000282414 Homo sapiens Species 0.000 description 1
- 241000187479 Mycobacterium tuberculosis Species 0.000 description 1
- 208000032023 Signs and Symptoms Diseases 0.000 description 1
- 230000009471 action Effects 0.000 description 1
- 230000001580 bacterial effect Effects 0.000 description 1
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- 229940079593 drug Drugs 0.000 description 1
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- 230000036541 health Effects 0.000 description 1
- 238000000338 in vitro Methods 0.000 description 1
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- 238000012360 testing method Methods 0.000 description 1
- 229960001005 tuberculin Drugs 0.000 description 1
- 229960002109 tuberculosis vaccine Drugs 0.000 description 1
- 238000002255 vaccination Methods 0.000 description 1
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Abstract
The invention discloses a group of tuberculosis serum exosome mi RNA markers and application thereof, and relates to the technical field of distributed power control devices, comprising the following components, S1 and markers; s2, an application system; s3, obtaining a Ct value: obtaining a Ct value obtained by performing real-time fluorescent quantitative reaction on a mi RNA marker in a sample; s4, contrast treatment: subtracting the Ct value from the internal reference, performing standardization processing to obtain a CT value, processing the CT value by using an SVM model, analyzing the risk probability value of the sample, and comparing the risk probability value with a threshold value to predict the risk of active tuberculosis. The invention adopts the provided 3 novel mi RNA markers related to active tuberculosis to prepare products, can directly carry out PCR detection, can synchronously analyze a plurality of mi RNA markers in the detection process, improves the detection rate, has simple and quick detection, does not cause false positive, and is more beneficial to comprehensively and accurately predicting the risk of active tuberculosis.
Description
Technical Field
The invention relates to the technical field of molecular diagnosis, in particular to a group of pulmonary tuberculosis serum exosome mi RNA markers and application thereof.
Background
Tuberculosis is a chronic pulmonary infectious disease caused by the infection of a complex group of mycobacterium tuberculosis, which seriously threatens the health of human beings. The high morbidity and mortality of tuberculosis, as well as the air-borne, place a considerable burden on the patient, the things the patient contacts, and the society. Therefore, it is especially important to find early diagnosis and early control of tuberculosis and also an important means for suppressing tuberculosis epidemic. The gold standard for the diagnosis of active tuberculosis and activity assessment after treatment is etiology examination. However, for patients with active tuberculosis with negative etiology examination, the judgment standard is lacking, and the final judgment can be made by combining various means such as clinical manifestation, treatment history, images and the like; and because of the unsatisfactory effect of tuberculosis vaccine, many patients with bacterial negative tuberculosis, especially patients without special symptoms and signs, cannot be effectively treated, the optimal treatment time for resisting tuberculosis is delayed, the risk of community transmission is increased, and tuberculin tests cannot distinguish natural tuberculosis infection from BCG vaccination. Therefore, a group of tuberculosis serum exosome mi RNA markers needs to be found, and a rapid, sensitive, efficient and stable diagnosis method is provided for tuberculosis diagnosis and differential diagnosis. The following problems exist in the prior art:
1. the existing tuberculosis serum exosome mi RNA markers and application thereof have defects in clinical diagnosis methods related to tuberculosis, have low detection sensitivity and are influenced by various factors;
2. the existing tuberculosis serum exosome mi RNA markers and application thereof have high false positive and false negative rates, and the sensitivity and specificity are limited at present.
Disclosure of Invention
In order to solve the technical problems, the invention adopts the following technical scheme:
a group of tuberculosis serum exosome mi RNA markers and application thereof, comprising the following contents,
s1, marker composition: exosome mi RNA biomarkers include one or more of has-miRNA-375-3p, has-miRNA-885-3P, has-miRNA-100-5 p;
s2, an application system: the mi RNA marker is a mi RNA marker associated with active tuberculosis, the system comprising: a data acquisition unit: for subjecting the sample to a real-time fluorescent quantitative reaction based on Direct S-Poly (T) Plus;
s3, obtaining a Ct value: obtaining a Ct value obtained by performing real-time fluorescent quantitative reaction on a mi RNA marker in a sample;
s4, contrast treatment: subtracting the Ct value from the internal reference, performing standardization processing to obtain a CT value, processing the CT value by using an SVM model, analyzing the risk probability value of the sample, and comparing the risk probability value with a threshold value to predict the risk of active tuberculosis.
The technical scheme of the invention is further improved as follows: s2 if two or more miRNA markers are increased in the sample from the subject, the possible risk of tuberculosis of the subject is indicated, the sample is a serum sample, and the kit comprises a reagent for detecting one miRNA in has-miRNA-375-3p and has-miRNA-885-3P, has-miRNA-100-5 p.
The technical scheme of the invention is further improved as follows: according to the primer designed by the mi RNA marker, the corresponding mi RNA marker can be detected rapidly according to the provided primer, nucleic acid extraction is not needed, samples are saved, and one mi RNA is detected to be at most not more than 1 mu L; and meanwhile, further analyzing according to the obtained result to obtain the result.
The technical scheme of the invention is further improved as follows: quantitative reverse transcription PCR or array hybridization was used to determine the level in has-miRNA-375-3p, has-miRNA-885-3P, has-miRNA-100-5 p.
The technical scheme of the invention is further improved as follows: providing a mi RNA antagonist.
The technical scheme of the invention is further improved as follows: the mi RNA antagonist is one of has-miRNA-375-3p and has-miRNA-885-3P, has-miRNA-100-5p antagonists.
By adopting the technical scheme, compared with the prior art, the invention has the following technical progress:
1. the invention provides a group of tuberculosis serum exosome mi RNA markers and application thereof, 3 new mi RNA markers related to active tuberculosis are formed by the markers, as the mi RNA markers have relatively stable content and good in vitro stability, the sensitivity is relatively strong in the detection process, the provided mi RNA markers are specific mi RNA markers for active tuberculosis diseases, the provided 3 mi RNA markers have specific expression profiles, and certain mi RNA can be used for predicting the occurrence of the diseases in early disease, so that early prediction experiments are more facilitated; in the detection and analysis process, the PCR detection can be directly carried out without screening and preparing specific antibodies, in the detection process, the synchronous analysis can be carried out on a plurality of mi RNA markers at the same time, the detection rate is improved, the detection is simple and rapid, false positives are not caused, the method is more beneficial to comprehensively and accurately predicting the risk of active tuberculosis, 3 novel mi RNA markers related to active tuberculosis are adopted for preparing products, the obtained products do not need to screen and prepare specific antibodies in the detection and analysis process, the PCR detection can be directly carried out, in the detection process, the synchronous analysis can be carried out on a plurality of mi RNA markers at the same time, the detection rate is improved, the detection is simple and rapid, false positives are not caused, and the method is more beneficial to comprehensively and accurately predicting the risk of active tuberculosis.
2. The invention provides a group of tuberculosis serum exosome mi RNA markers and application thereof, and the kit comprises a reagent for detecting mi RNA in has-miRNA-375-3p and has-miRNA-885-3P, has-miRNA-100-5p under the action of the kit of an application system, so that the quantitative determination is accurate, the detection result is stable and reliable, and the sensitivity and the specificity of clinical early diagnosis can be improved. The kit combines a common RNA extraction reagent and a common reverse transcription reagent, and can rapidly detect the corresponding mi RNA marker according to the primer designed by the mi RNA marker, so that nucleic acid extraction is not needed, the pollution is reduced, the loss is avoided, the cost is reduced, and the technical threshold is low; sample saving, detecting a mi RNA of at most 1 mu L sample; meanwhile, the method can further analyze according to the obtained result, the obtained result is more reliable, the method can be widely used, the Ct value of the mi RNA marker of the sample obtained by real-time fluorescence quantitative reaction analysis is firstly standardized, the error range is ensured to be smaller, and the result is more reliable; and then analyzing the risk probability value of the sample by using the SVM model and comparing the risk probability value with a threshold value, combining the real-time fluorescence quantitative reaction with an AI diagnosis model, accurately and rapidly predicting the risk of active tuberculosis, and having higher accuracy and higher reliability of a prediction result, and being widely applied.
Drawings
FIG. 1 is a schematic flow chart of a group of tuberculosis serum exosome mi RNA markers and application thereof.
Detailed Description
The invention is further illustrated by the following examples:
example 1
As shown in FIG. 1, the invention provides a group of tuberculosis serum exosome mi RNA markers and application thereof, comprising the following contents,
s1, marker composition: exosome mi RNA biomarkers include one or more of has-miRNA-375-3p, has-miRNA-885-3P, has-miRNA-100-5 p;
s2, an application system: the mi RNA marker is a mi RNA marker associated with active tuberculosis, the system comprising: a data acquisition unit: for subjecting the sample to a real-time fluorescent quantitative reaction based on Direct S-Poly (T) Plus;
s3, obtaining a Ct value: obtaining a Ct value obtained by performing real-time fluorescent quantitative reaction on a mi RNA marker in a sample;
s4, contrast treatment: subtracting the Ct value from the internal reference, performing standardization processing to obtain a CT value, processing the CT value by using an SVM model, analyzing the risk probability value of the sample, and comparing the risk probability value with a threshold value to predict the risk of active tuberculosis.
In the embodiment, the provided 3 mi RNA markers have specific expression profiles, and certain mi RNAs can be used for predicting the occurrence of diseases in early stages of the diseases, so that early-stage prediction experiments are facilitated; in the detection and analysis process, the PCR detection can be directly carried out without screening and preparing specific antibodies, in the detection process, the synchronous analysis can be carried out on a plurality of mi RNA markers at the same time, the detection rate is improved, the detection is simple and rapid, false positives are not caused, the method is more beneficial to comprehensively and accurately predicting the risk of active tuberculosis, 3 novel mi RNA markers related to active tuberculosis are adopted for preparing products, the obtained products do not need to screen and prepare specific antibodies in the detection and analysis process, the PCR detection can be directly carried out, in the detection process, the synchronous analysis can be carried out on a plurality of mi RNA markers at the same time, the detection rate is improved, the detection is simple and rapid, false positives are not caused, and the method is more beneficial to comprehensively and accurately predicting the risk of active tuberculosis.
Example 2
As shown in fig. 1, on the basis of embodiment 1, the present invention provides a technical solution: preferably, if two or more mi RNA markers are raised in the S2 sample from the subject, the risk that the subject may have tuberculosis is indicated, the sample is a serum sample, the kit comprises reagents for detecting one mi RNA in has-miRNA-375-3p, has-miRNA-885-3P, has-miRNA-100-5p, the corresponding mi RNA marker can be rapidly detected according to the primer designed according to the mi RNA marker, the nucleic acid is not required to be extracted, the sample is saved, and the maximum one mi RNA is not more than 1 μl sample; and meanwhile, further analyzing according to the obtained result to obtain a result, and determining the level of the has-miRNA-375-3p and the has-miRNA-885-3P, has-miRNA-100-5p by utilizing quantitative reverse transcription PCR or array hybridization to provide the application of the mi RNA antagonist in preparing the medicines for preventing or alleviating the subjects, wherein the mi RNA antagonist is one of the has-miRNA-375-3p and the has-miRNA-885-3P, has-miRNA-100-5p antagonist.
In this embodiment, the sensitivity and specificity of the clinical early diagnosis can be improved. The kit combines a common RNA extraction reagent and a common reverse transcription reagent, and can rapidly detect the corresponding mi RNA marker according to the primer designed by the mi RNA marker, so that nucleic acid extraction is not needed, the pollution is reduced, the loss is avoided, the cost is reduced, and the technical threshold is low; sample saving, detecting a mi RNA of at most 1 mu L sample; meanwhile, the method can further analyze according to the obtained result, the obtained result is more reliable, the method can be widely used, the Ct value of the mi RNA marker of the sample obtained by real-time fluorescence quantitative reaction analysis is firstly standardized, the error range is ensured to be smaller, and the result is more reliable; and then analyzing the risk probability value of the sample by using the SVM model and comparing the risk probability value with a threshold value, combining the real-time fluorescence quantitative reaction with an AI diagnosis model, accurately and rapidly predicting the risk of active tuberculosis, and having higher accuracy and higher reliability of a prediction result, and being widely applied.
The working principle of the group of tuberculosis serum exosome mi RNA markers and the application thereof is specifically described below.
As shown in fig. 1, the marker composition exosome mi RNA biomarker comprises one or more of has-miRNA-375-3p, has-miRNA-885-3P, has-miRNA-100-5p, 3 mi RNA markers have specific expression profiles and certain mi RNAs can be used for predicting the occurrence of diseases in early stages of the diseases, which is more beneficial for performing early prediction experiments; in the detection and analysis process, the PCR detection can be directly carried out without screening and preparing specific antibodies, in the detection process, the synchronous analysis can be carried out on a plurality of mi RNA markers at the same time, the detection rate is improved, the detection is simple and rapid, false positives are not caused, the method is more beneficial to comprehensively and accurately predicting the risk of active tuberculosis, 3 novel mi RNA markers related to active tuberculosis are adopted for preparing products, the obtained products do not need to screen and prepare specific antibodies in the detection and analysis process, the PCR detection can be directly carried out, in the detection process, the synchronous analysis can be carried out on a plurality of mi RNA markers at the same time, the detection rate is improved, the detection is simple and rapid, false positives are not caused, the method is more beneficial to comprehensively and accurately predicting the risk of active tuberculosis, and the mi RNA markers are mi RNA markers related to active tuberculosis, and the system comprises: a data acquisition unit: the kit is used for carrying out a real-time fluorescence quantitative reaction based on Direct S-Poly (T) Plus, if the level of a mi RNA marker is increased in a sample from a subject, the condition that the subject has rejection reaction or has the risk of having rejection reaction is indicated, the sample is a serum sample, the kit comprises reagents for detecting at least one mi RNA in has-miRNA-375-3p and has-miRNA-885-3P, has-miRNA-100-5p, the corresponding mi RNA marker can be rapidly detected according to the primer designed according to the mi RNA marker, nucleic acid is not required to be extracted, the sample is saved, and the maximum one mi RNA is not more than 1 mu L of sample; meanwhile, the method further analyzes according to the obtained result to obtain the result, and the quantitative reverse transcription PCR or array hybridization is utilized to determine the levels of has-miRNA-375-3p and has-miRNA-885-3P, has-miRNA-100-5p, and the kit is combined with a common RNA extraction reagent and a common reverse transcription reagent, so that the primer designed according to the mi RNA marker can rapidly detect the corresponding mi RNA marker without extracting nucleic acid, and is simple and rapid, pollution-reducing, loss-avoiding, cost-reducing and low in technical threshold; sample saving, detecting a mi RNA of at most 1 mu L sample; meanwhile, the method can further analyze according to the obtained result, the obtained result is more reliable, the method can be widely used, the Ct value of the mi RNA marker of the sample obtained by real-time fluorescence quantitative reaction analysis is firstly standardized, the error range is ensured to be smaller, and the result is more reliable; and analyzing the risk probability value of the sample by using the SVM model, comparing the risk probability value with a threshold value, combining the real-time fluorescent quantitative reaction with an AI diagnosis model, accurately and rapidly predicting the active tuberculosis disease risk, wherein the accuracy of the prediction result is higher, the reliability is higher, the method can be widely applied, the method can be used for obtaining the Ct value obtained by carrying out the real-time fluorescent quantitative reaction on the mi RNA marker in the sample, subtracting the Ct value from an internal reference, carrying out standardized processing to obtain the CT value, processing the CT value by using the SVM model, analyzing the risk probability value of the sample, and comparing the risk probability value with the threshold value to predict the active tuberculosis disease risk.
The foregoing invention has been generally described in great detail, but it will be apparent to those skilled in the art that modifications and improvements can be made thereto. Accordingly, it is intended to cover modifications or improvements within the spirit of the inventive concepts.
Claims (6)
1. A group of tuberculosis serum exosome mi RNA markers and application thereof, comprising the following contents, and characterized in that:
s1, marker composition: exosome mi RNA biomarkers include one or more of has-miRNA-375-3p, has-miRNA-885-3P, has-miRNA-100-5 p;
s2, an application system: the mi RNA marker is a mi RNA marker associated with active tuberculosis, the system comprising: a data acquisition unit: for subjecting the sample to a real-time fluorescent quantitative reaction based on Direct S-Poly (T) Plus;
s3, obtaining a Ct value: obtaining a Ct value obtained by performing real-time fluorescent quantitative reaction on a mi RNA marker in a sample;
s4, contrast treatment: subtracting the Ct value from the internal reference, performing standardization processing to obtain a CT value, processing the CT value by using an SVM model, analyzing the risk probability value of the sample, and comparing the risk probability value with a threshold value to predict the risk of active tuberculosis.
2. A set of tuberculosis serum exosomes mi RNA markers-markers as described in claim 1, and their use, characterized in that: s2 if two or more miRNA markers are increased in the sample from the subject, the risk of tuberculosis of the subject is indicated, the sample is a serum sample, and the kit comprises a reagent for detecting one miRNA in has-miRNA-375-3p and has-miRNA-885-3P, has-miRNA-100-5 p.
3. A set of tuberculosis serum exosomes mi RNA markers-markers as in claim 2, and their use, characterized in that: according to the primer designed by the mi RNA marker, the corresponding mi RNA marker can be detected rapidly according to the provided primer, nucleic acid extraction is not needed, samples are saved, and one mi RNA is detected to be at most not more than 1 mu L; and meanwhile, further analyzing according to the obtained result to obtain the result.
4. A set of tuberculosis serum exosomes mi RNA markers-markers as in claim 2, and their use, characterized in that: quantitative reverse transcription PCR or array hybridization was used to determine the level in has-miRNA-375-3p, has-miRNA-885-3P, has-miRNA-100-5 p.
5. A set of tuberculosis serum exosomes mi RNA markers-markers as in claim 2, and their use, characterized in that: providing a mi RNA antagonist.
6. The set of tuberculosis serum exosomes mi RNA markers-set according to claim 5, and their use, characterized in that: the mi RNA antagonist is one of a has-miRNA-375-3P antagonist, a has-miRNA-885-3P antagonist and a has-miRNA-100-5P antagonist.
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| CN117637027B (en) * | 2024-01-26 | 2024-03-29 | 南通大学附属医院 | Efficiency evaluation method for diagnosing early lung adenocarcinoma by exosome circRNAs |
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