CN109097473A - A kind of non-small cell lung cancer auxiliary diagnostic box - Google Patents
A kind of non-small cell lung cancer auxiliary diagnostic box Download PDFInfo
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- CN109097473A CN109097473A CN201810974389.8A CN201810974389A CN109097473A CN 109097473 A CN109097473 A CN 109097473A CN 201810974389 A CN201810974389 A CN 201810974389A CN 109097473 A CN109097473 A CN 109097473A
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- lung cancer
- small cell
- cell lung
- primer
- auxiliary diagnostic
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- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
- C12Q1/00—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions
- C12Q1/68—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving nucleic acids
- C12Q1/6876—Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes
- 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
- C12Q1/6886—Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes for diseases caused by alterations of genetic material for cancer
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- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
- C12Q2600/00—Oligonucleotides characterized by their use
- C12Q2600/158—Expression markers
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- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
- C12Q2600/00—Oligonucleotides characterized by their use
- C12Q2600/178—Oligonucleotides characterized by their use miRNA, siRNA or ncRNA
Abstract
The invention discloses a kind of non-small cell lung cancer auxiliary diagnostic boxes.MiRNA647 is preparing the application in non-small cell lung cancer auxiliary diagnostic as the miRNA marker of detection non-small cell lung cancer.A kind of non-small cell lung cancer auxiliary diagnostic box, including downstream primer shown in the upstream primer as shown in SEQ ID NO.1 and SEQ ID NO.2.It is related with the generation of non-small cell lung cancer that present invention finds miRNA647 expressions, pass through the expression of miRNA647 in detection blood, it may determine that whether it suffers from non-small cell lung cancer perhaps with the presence or absence of the risk with non-small cell lung cancer so that clinician be instructed to give its prevention scheme or therapeutic scheme in conjunction with clinical symptoms.Therefore, detecting the reagents such as primer, the probe of miRNA647 expression can be used for preparing non-small cell lung cancer auxiliary diagnostic.
Description
Technical field
The invention belongs to biological medicine detection fields, are related to a kind of non-small cell lung cancer auxiliary diagnostic box.
Background technique
Lung cancer is the malignant tumour of incidence and case fatality rate first in current world wide, annual new hair patient about 50
Ten thousand, although having many progress in pulmonary cancer diagnosis treatment in recent years, survival rate is only 15%-17% within 5 years.From tissue
On, lung cancer is divided into Small Cell Lung Cancer (small cell lung cancer, SCLC) and non-small cell lung cancer (non-small
Cell lung cancer, NSCLC), proportion is respectively 15% and 85%.Wherein non-small cell lung cancer is mostly late
It can be found, 5 years survival rates are less than 15%, so survival rate will increase if can accomplish to early diagnose.It is existing
The main means of the detection in stage are iconography and fiberoptic bronchoscopy, and compared with the two, tumor-marker analyte detection has
The advantages that risk is small, radiationless, inexpensive and easy.
MicroRNA (miRNA) is the single-stranded microRNA of endogenous that a kind of size is 19-22 nucleotide, is deposited extensively
It is in eucaryote, it is highly conserved during biological evolution, it can be specific expressed in cell.MiRNA can pass through
Silencing complex (RISC) is guided with target mRNA specific base complementary pairing, degrades and mRNA or checks its translation, transcribing
Level tune related gene expression afterwards, to play an important role in processes such as cell Proliferation, differentiation, apoptosis.Studies have shown that
MiRNA is closely related with tumour, and the missing or amplification of chromosome are frequently accompanied by the generation or development process of tumour, from
And lead to the activation of oncogene or the inactivation of tumor suppressor gene.Studies have shown that many miRNA molecules are located in and the close phase of tumour
In the chromosomal variation area of pass, single miRNA can target multiple mRNA, so that the expression of multiple albumen is adjusted, miRNA table
It is an important factor for leading to a series of formation of tumours up to disorder.MiRNA becomes research hotspot in recent years, research shows that miRNA is logical
The expression for crossing regulation target gene can influence the various procedures such as proliferation, invasion, transfer, the apoptosis of tumour.For example, miRNA-124
It can inhibit the proliferation of breast cancer cell by lowering the expression of CDK4；MiRNA-214 makes breast cancer cell by acting on p53
Invasive ability enhancing；MiRNA-144 promotes Increase Apoptosis of Lung Cancer Cells.The unconventionality expression of miRNA can induce and disease dependency basis
The expression of cause is abnormal, thus the generation of induced tumor, therefore the expression of directly detection miRNA can facing for tumour
Bed diagnosis provides certain reference, plays it emphatically in terms of the clinical diagnosis of various tumor diseases, treatment and prognosis evaluation
It acts on.
Summary of the invention
The purpose of the present invention is in view of the above drawbacks of the prior art, provide a kind of miRNA for detecting non-small cell lung cancer
The application of marker.
It is a further object of the present invention to provide the primer, probe or the genetic chips that detect the marker to prepare non-small cell
Application in lung cancer auxiliary diagnostic.
It is yet another object of the invention to provide a kind of non-small cell lung cancer auxiliary diagnostic boxes.
The purpose of the present invention can be achieved through the following technical solutions:
MiRNA647 is preparing the examination of non-small cell lung cancer auxiliary diagnosis as the miRNA marker of detection non-small cell lung cancer
Application in agent.
Primer, probe or the genetic chip of detection miRNA647 is preparing answering in non-small cell lung cancer auxiliary diagnostic
With.
The primer preferably upstream primer as shown in SEQ ID NO.1 and SEQ ID NO.2 of the detection miRNA647
Shown in downstream primer composition.
A kind of primer being used to prepare non-small cell lung cancer auxiliary diagnostic, it is characterised in that by SEQ ID NO.1 institute
The composition of downstream primer shown in the upstream primer and SEQ ID NO.2 shown.
A kind of non-small cell lung cancer auxiliary diagnostic box, including above-mentioned primer.
The kit, it is also preferable to include the primer of detection reference gene U6 and other conventional reagents of qPCR.
The utility model has the advantages that
It is related with the generation of non-small cell lung cancer that present invention finds miRNA647 expressions, by detection blood
The expression of miRNA647 may determine that whether it suffers from non-small cell lung cancer or with the presence or absence of suffering from conjunction with clinical symptoms
The risk of non-small cell lung cancer, so that clinician be instructed to give its prevention scheme or therapeutic scheme.Therefore, it detects
The reagents such as primer, the probe of miRNA647 expression can be used for preparing non-small cell lung cancer auxiliary diagnostic.
Detailed description of the invention
The relative expression quantity of Fig. 1 miRNA-647
Specific embodiment
Embodiment 1
1, RNA is extracted
By the blood sample of the non-small cell lung cancer of collection and Healthy People each 35, PAXgene Blood RNA is used
Kit (Qiagen, Gemany) extracts the RNA of blood, and all steps are operated to specifications, after RNA is extracted, 1 μ L is taken to use
1% agarose gel electrophoresis detection, and with Nanodrop2000 (Thermo Scientific) detectable concentration, when 260/280
When ratio is 2.0, RNA purity is 100%, and between 1.8-2.0, RNA purity can be used 260/280 ratio close to 80-90%
In reverse transcription.
2, the design of primer and probe
The gene order that miRNA-647 and internal reference U6 are learnt according to miRBse inquiry, is designed using software Primer 5.0
The amplimer of microRNA.
3, the expression of real-time fluorescence quantitative PCR detection miRNA-647
(1) it reaction system: see the table below
(2) it reaction condition: see the table below
(4) statistical analysis
According to obtained Ct value, pass through 2-△△CtMethod calculate miRNA-647 using U6 as the relative expression levels of internal reference
Numerical value.
The Ct value obtained according to real-time fluorescence quantitative PCR result, passes through 2-△△CtMethod calculate miRNA-647 with U6
For the numerical value of the relative expression levels of internal reference.The result shows that as shown in Figure 1, compared with the blood of Healthy People, non-small cell lung cancer
The expression of miRNA-647 significantly increases in blood.
2. sensitivity technique of embodiment
The cDNA that reverse transcription obtains is diluted according to 10 multiple proportions, is finally diluted to 107Times, take different dilution ratios
CDNA do quantitative fluorescent PCR according to the method for embodiment 1, detection miRNA-647 can detect the maximum dilution multiple of cq value.
From following table as can be seen that miRNA-647 is up to being diluted to 106It just can't detect cq value again.According to 100% reverse
Efficiency similitude is recorded, the detection of cDNA detection miRNA-647 is limited to 3.5 × 10-2Ng/ μ L illustrates that cDNA sensitivity is higher.
Sequence table
<110>Nanjing Qiu Zhen Gene Tech. Company Limited
<120>a kind of non-small cell lung cancer auxiliary diagnostic box
<160> 4
<170> SIPOSequenceListing 1.0
<210> 1
<211> 18
<212> DNA
<213>artificial sequence (Artificial Sequence)
<400> 1
tggctgcact cacttcct 18
<210> 2
<211> 18
<212> DNA
<213>artificial sequence (Artificial Sequence)
<400> 2
aaggaagtga gtgcagcc 18
<210> 3
<211> 20
<212> DNA
<213>artificial sequence (Artificial Sequence)
<400> 3
tgcgggtgct cgcttcggca 20
<210> 4
<211> 20
<212> DNA
<213>artificial sequence (Artificial Sequence)
<400> 4
aacgcttcac gaatttgcgt 20
Claims (6)
1.miRNA647 is preparing non-small cell lung cancer auxiliary diagnostic as the miRNA marker of detection non-small cell lung cancer
In application.
2. primer, probe or the genetic chip of detection miRNA647 are preparing answering in non-small cell lung cancer auxiliary diagnostic
With.
3. application according to claim 2, it is characterised in that the primer of the detection miRNA647 is by SEQ ID NO.1
Shown in downstream primer composition shown in upstream primer and SEQ ID NO.2.
4. a kind of primer for being used to prepare non-small cell lung cancer auxiliary diagnostic, it is characterised in that as shown in SEQ ID NO.1
Upstream primer and SEQ ID NO.2 shown in downstream primer composition.
5. a kind of non-small cell lung cancer auxiliary diagnostic box, it is characterised in that including primer as claimed in claim 4.
6. kit according to claim 5, it is characterised in that further include the primer and qPCR for detecting reference gene U6
Other conventional reagents.
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| Application Number | Priority Date | Filing Date | Title |
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| CN201810974389.8A CN109097473A (en) | 2018-08-24 | 2018-08-24 | A kind of non-small cell lung cancer auxiliary diagnostic box |
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| CN201810974389.8A CN109097473A (en) | 2018-08-24 | 2018-08-24 | A kind of non-small cell lung cancer auxiliary diagnostic box |
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Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN111349705A (en) * | 2020-03-18 | 2020-06-30 | 昆明医科大学 | CircASXL1 as lung cancer diagnosis marker and application thereof |
| CN111424093A (en) * | 2020-03-30 | 2020-07-17 | 中国医学科学院肿瘤医院 | Kit, device and method for lung cancer diagnosis |
Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN105308189A (en) * | 2013-04-15 | 2016-02-03 | 瑞泽恩制药公司 | Markers of tumor cell response to anti-cancer therapy |
| CN106974929A (en) * | 2017-03-24 | 2017-07-25 | 南方医科大学南方医院 | Purposes of the miR 647 in the medicine for preparing treatment stomach cancer |
-
2018
- 2018-08-24 CN CN201810974389.8A patent/CN109097473A/en active Pending
Patent Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN105308189A (en) * | 2013-04-15 | 2016-02-03 | 瑞泽恩制药公司 | Markers of tumor cell response to anti-cancer therapy |
| CN106974929A (en) * | 2017-03-24 | 2017-07-25 | 南方医科大学南方医院 | Purposes of the miR 647 in the medicine for preparing treatment stomach cancer |
Non-Patent Citations (1)
| Title |
|---|
| SANTOSH K. PATNAIK ET AL: "Evaluation of MicroRNA Expression Profiles That May Predict Recurrence of Localized Stage I Non–Small Cell Lung Cancer after Surgical Resection", 《CANCER RES》 * |
Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN111349705A (en) * | 2020-03-18 | 2020-06-30 | 昆明医科大学 | CircASXL1 as lung cancer diagnosis marker and application thereof |
| CN111424093A (en) * | 2020-03-30 | 2020-07-17 | 中国医学科学院肿瘤医院 | Kit, device and method for lung cancer diagnosis |
| CN111424093B (en) * | 2020-03-30 | 2021-10-15 | 中国医学科学院肿瘤医院 | Kit, device and method for lung cancer diagnosis |
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Application publication date: 20181228 |