CN108796073A - A kind of kit of joint-detection gene maspin and miR-17 Diagnosis of Breast cancer - Google Patents
A kind of kit of joint-detection gene maspin and miR-17 Diagnosis of Breast cancer Download PDFInfo
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- CN108796073A CN108796073A CN201810416130.1A CN201810416130A CN108796073A CN 108796073 A CN108796073 A CN 108796073A CN 201810416130 A CN201810416130 A CN 201810416130A CN 108796073 A CN108796073 A CN 108796073A
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- C12Q1/6883—Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes for diseases caused by alterations of genetic material
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Abstract
The invention discloses a kind of kits of joint-detection gene maspin and miR-17 Diagnosis of Breast cancer, belong to medical diagnosis on disease field.The present invention has found that maspin is negatively correlated with breast cancer by research, and miR-17 is proportionate with breast cancer, the two gene pairs breast cancer of joint-detection, which carry out diagnosis, very high accuracy rate.The kit of the present invention includes the primer of detection gene maspin and miR-17 expression, and the primer of detection reference gene expression, also includes real-time quantitative PCR reaction reagent；The sequence of the primer is shown in SEQ ID NO.1-8.Be detected using kit of the present invention, when maspin Ct values more than 31, miR-17 Ct values be less than 12 when, can tentative diagnosis be breast cancer.Present invention Diagnosis of Breast cancer from gene level is more accurate than the tissue of traditional extraction detected person and alleviates the pain of patient.
Description
Technical field
The present invention relates to medical diagnosis on disease fields, and in particular to a kind of joint-detection gene maspin and miR-17 Diagnosis of Breast
The kit of cancer.
Background technology
Breast cancer has become the kinds of tumor for threatening women physical and mental health, and breast cancer survival rate and discovery have very high point sooner or later
System, traditional diagnostic method have：Breast cancer Mo-target-X-ray photography, B ultrasound scanning, the swollen object Puncture cytology inspection of fine needle etc..These sides
Method detection time is longer, and larger to the somatic damage of detected person, and the purpose of early screening is not achieved.
The present invention has found the expression of joint-detection maspin and miR-17 in serum gene level, energy by studying
Whether enough accurate judgement testers suffer from breast cancer.
Invention content
It is an object of the invention to be directed to the problems of existing breast cancer conventional method, a kind of joint-detection base is provided
Because of the kit of maspin and miR-17 Diagnosis of Breast cancers.
The purpose of the invention is achieved by the following technical solution：
The present invention has found that maspin is negatively correlated with breast cancer by research, and miR-17 is proportionate with breast cancer, joint
Detecting the two gene pairs breast cancer and diagnose has very high accuracy rate, and therefore, gene maspin and miR-17, which have, to be prepared
The purposes of Diagnosis of Breast cancer kit.
A kind of kit of joint-detection gene maspin and miR-17 Diagnosis of Breast cancer, including detection gene maspin with
The primer of miR-17 expression, and detect the primer of reference gene expression.When maspin is target gene, GAPDH is internal reference；
When miR-17 is target gene, U6 is internal reference.
It is further preferred that the primer sequence of the above-mentioned each gene of detection is as follows：
Maspin sense primers：TGTAAATAAAGGGATGG,
Maspin downstream primers：GATGTGGAGGATGAGTC；
MiR-17 sense primers：TGTCAAAGTGCTTACAGTG,
MiR-17 downstream primers：CACAGCTCGTAGAACAGGAGG；
GAPDH sense primers：CGGAGTCAACGGATTTGGTCGTAT,
GAPDH downstream primers：AGCCTTCTCCATGGTGGTGAAGAC；
U6 sense primers：CTCGCTTCGGCAGCACA,
U6 downstream primers：AACGCTTCACGAATTTGCGT.
It is further preferred that the kit of joint-detection gene maspin and miR-17 the Diagnosis of Breast cancer, is also wrapped
Reaction reagent containing real-time quantitative PCR.
It carries out reverse transcription reaction by extracting the RNA in tester's blood sample using kit of the present invention and obtains
CDNA, then real-time quantitative PCR reaction is carried out by template of cDNA, inspection may determine that according to the expression of maspin and miR-17
Whether survey person suffers from breast cancer.
The present invention has the following advantages compared with the prior art and advantageous effect：Present invention finds maspin and breast cancer
Negatively correlated, miR-17 is proportionate with breast cancer.It is detected using kit of the present invention, when the Ct values of maspin are more than
31, miR-17 Ct values be less than 12 when, can tentative diagnosis be breast cancer.Present invention Diagnosis of Breast cancer ratio tradition from gene level
Extraction detected person tissue it is more accurate and alleviate the pain of patient.
Specific implementation mode
Following embodiment should not be construed as limiting the invention for further illustrating the present invention.If not referring in particular to
Conventional means bright, that technological means used in embodiment is well known to those skilled in the art.
Embodiment 1
A kind of kit of joint-detection gene maspin and miR-17 Diagnosis of Breast cancer, including following primer：
Maspin sense primers：TGTAAATAAAGGGATGG,
Maspin downstream primers：GATGTGGAGGATGAGTC；
MiR-17 sense primers：TGTCAAAGTGCTTACAGTG,
MiR-17 downstream primers：CACAGCTCGTAGAACAGGAGG；
GAPDH sense primers：CGGAGTCAACGGATTTGGTCGTAT,
GAPDH downstream primers：AGCCTTCTCCATGGTGGTGAAGAC；
U6 sense primers：CTCGCTTCGGCAGCACA,
U6 downstream primers：AACGCTTCACGAATTTGCGT.
Embodiment 2
The method being detected using the kit described in embodiment 1, is included the following steps：
(1) RNA in tester's blood sample is extracted.
(2) reverse transcription reagent box for using TaKaRa companies carries out reverse transcription reaction and obtains cDNA.
(3) real-time quantitative PCR reaction is carried out by template of cDNA, reaction system and reaction condition are as follows.
Reaction system：SYBR Premix Ex Taq (2 ×) 12.5 μ L, sense primer (10 μM) 1 μ L, downstream primer (10 μ
M) 1 μ L, template (cDNA solution) 0.5 μ L, ddH2O 10μL。
Reaction condition：It is acted on 3 minutes with 95 DEG C；PCR carry out 35 cycle, denaturation 95 DEG C 30 seconds, annealing 58 DEG C 30 seconds, prolong
Stretch 72 DEG C 30 seconds.
10 groups of non-breast cancer patients and patient with breast cancer are had detected according to the method described above, and real-time quantitative PCR concrete outcome is：
(Cycle threshold, the fluorescence signal in each reaction tube reach the Ct values of maspin in 10 patient with breast cancer's blood
The recurring number undergone when the thresholding of setting) it is more than 31；The Ct values of miR-17 in 10 patient with breast cancer's blood are less than 12.It is logical
It crosses detection and finds that the expression quantity of non-breast cancer patient maspin is higher than the expression quantity of patient with breast cancer；Non-breast cancer patient miR-17
Expression quantity it is lower than the expression quantity in patient with breast cancer blood, show that maspin is negatively correlated with breast cancer, miR-17 and mammary gland
Cancer is proportionate.
100 patient with breast cancers are randomly choosed again, detect maspin in these patient with breast cancer's blood according to the method described above
With the expression of miR-17, wherein the Ct values of 95 patient with breast cancer maspin be more than 31,100 patient with breast cancer miR-
17Ct values are less than 12, should be the result shows that Detection accuracy of the present invention is high.
The above embodiment is a preferred embodiment of the present invention, but embodiments of the present invention are not by above-described embodiment
Limitation, it is other it is any without departing from the spirit and principles of the present invention made by changes, modifications, substitutions, combinations, simplifications,
Equivalent substitute mode is should be, is included within the scope of the present invention.
Sequence table
<110>Wuhan University Of Technology
<120>A kind of kit of joint-detection gene maspin and miR-17 Diagnosis of Breast cancer
<160> 8
<170> SIPOSequenceListing 1.0
<210> 1
<211> 17
<212> DNA
<213>Artificial sequence (Artificial Sequence)
<400> 1
tgtaaataaa gggatgg 17
<210> 2
<211> 17
<212> DNA
<213>Artificial sequence (Artificial Sequence)
<400> 2
gatgtggagg atgagtc 17
<210> 3
<211> 19
<212> DNA
<213>Artificial sequence (Artificial Sequence)
<400> 3
tgtcaaagtg cttacagtg 19
<210> 4
<211> 21
<212> DNA
<213>Artificial sequence (Artificial Sequence)
<400> 4
cacagctcgt agaacaggag g 21
<210> 5
<211> 24
<212> DNA
<213>Artificial sequence (Artificial Sequence)
<400> 5
cggagtcaac ggatttggtc gtat 24
<210> 6
<211> 24
<212> DNA
<213>Artificial sequence (Artificial Sequence)
<400> 6
agccttctcc atggtggtga agac 24
<210> 7
<211> 17
<212> DNA
<213>Artificial sequence (Artificial Sequence)
<400> 7
ctcgcttcgg cagcaca 17
<210> 8
<211> 20
<212> DNA
<213>Artificial sequence (Artificial Sequence)
<400> 8
aacgcttcac gaatttgcgt 20
Claims (5)
1. purposes of the gene maspin and miR-17 in preparing Diagnosis of Breast cancer kit.
2. a kind of kit of joint-detection gene maspin and miR-17 Diagnosis of Breast cancer, it is characterised in that：Including detection base
Because of the primer of maspin and miR-17 expression, and the primer of detection reference gene expression.
3. kit according to claim 2, it is characterised in that：When maspin is target gene, GAPDH is internal reference；
When miR-17 is target gene, U6 is internal reference.
4. kit according to claim 2, it is characterised in that：The primer sequence for detecting each gene is as follows：
Maspin sense primers：TGTAAATAAAGGGATGG,
Maspin downstream primers：GATGTGGAGGATGAGTC；
MiR-17 sense primers：TGTCAAAGTGCTTACAGTG,
MiR-17 downstream primers：CACAGCTCGTAGAACAGGAGG；
GAPDH sense primers：CGGAGTCAACGGATTTGGTCGTAT,
GAPDH downstream primers：AGCCTTCTCCATGGTGGTGAAGAC；
U6 sense primers：CTCGCTTCGGCAGCACA,
U6 downstream primers：AACGCTTCACGAATTTGCGT.
5. kit according to claim 2, it is characterised in that：Including real-time quantitative PCR reaction reagent.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201810416130.1A CN108796073A (en) | 2018-05-03 | 2018-05-03 | A kind of kit of joint-detection gene maspin and miR-17 Diagnosis of Breast cancer |
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| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201810416130.1A CN108796073A (en) | 2018-05-03 | 2018-05-03 | A kind of kit of joint-detection gene maspin and miR-17 Diagnosis of Breast cancer |
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| Publication Number | Publication Date |
|---|---|
| CN108796073A true CN108796073A (en) | 2018-11-13 |
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| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN201810416130.1A Pending CN108796073A (en) | 2018-05-03 | 2018-05-03 | A kind of kit of joint-detection gene maspin and miR-17 Diagnosis of Breast cancer |
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Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN111363819A (en) * | 2020-01-08 | 2020-07-03 | 武汉科技大学 | Method for jointly detecting and diagnosing breast cancer by utilizing ddPCR technology |
Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101201357A (en) * | 2007-11-05 | 2008-06-18 | 广州益善生物技术有限公司 | Liquid phase chip reagent box for early diagnosing mammary cancer and preparation method thereof |
| CN101993937A (en) * | 2009-08-10 | 2011-03-30 | 芮屈生物技术（上海）有限公司 | Maspin gene in-situ hybridization detection kit and detection method and use thereof |
| CN104630377A (en) * | 2015-02-28 | 2015-05-20 | 上海赛安生物医药科技有限公司 | Breast cancer miRNA detection kit and application of miRNA thereof |
| CN104991065A (en) * | 2015-07-07 | 2015-10-21 | 复旦大学附属金山医院 | Triple marker used for diagnosing breast cancer and application thereof |
-
2018
- 2018-05-03 CN CN201810416130.1A patent/CN108796073A/en active Pending
Patent Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101201357A (en) * | 2007-11-05 | 2008-06-18 | 广州益善生物技术有限公司 | Liquid phase chip reagent box for early diagnosing mammary cancer and preparation method thereof |
| CN101993937A (en) * | 2009-08-10 | 2011-03-30 | 芮屈生物技术（上海）有限公司 | Maspin gene in-situ hybridization detection kit and detection method and use thereof |
| CN104630377A (en) * | 2015-02-28 | 2015-05-20 | 上海赛安生物医药科技有限公司 | Breast cancer miRNA detection kit and application of miRNA thereof |
| CN104991065A (en) * | 2015-07-07 | 2015-10-21 | 复旦大学附属金山医院 | Triple marker used for diagnosing breast cancer and application thereof |
Non-Patent Citations (2)
| Title |
|---|
| FANGLIANG YANG等: "miR-17 as a diagnostic biomarker regulates cell proliferation in breast cancer", 《ONCOTARGETS AND THERAPY》 * |
| 朱文宇等: "miR-17在乳腺癌组织中表达与临床病理特征的相关性", 《中国保健营养》 * |
Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN111363819A (en) * | 2020-01-08 | 2020-07-03 | 武汉科技大学 | Method for jointly detecting and diagnosing breast cancer by utilizing ddPCR technology |
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Application publication date: 20181113 |