CN108841961A - Application of the LINC01702 in the diagnostic kit of preparation hepatocellular carcinoma - Google Patents
Application of the LINC01702 in the diagnostic kit of preparation hepatocellular carcinoma Download PDFInfo
- Publication number
- CN108841961A CN108841961A CN201810833682.2A CN201810833682A CN108841961A CN 108841961 A CN108841961 A CN 108841961A CN 201810833682 A CN201810833682 A CN 201810833682A CN 108841961 A CN108841961 A CN 108841961A
- Authority
- CN
- China
- Prior art keywords
- long
- linc01702
- hepatocellular carcinoma
- coding rna
- chain non
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Granted
Links
Classifications
-
- 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
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K45/00—Medicinal preparations containing active ingredients not provided for in groups A61K31/00 - A61K41/00
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P35/00—Antineoplastic agents
-
- 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
Landscapes
- Health & Medical Sciences (AREA)
- Chemical & Material Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Organic Chemistry (AREA)
- General Health & Medical Sciences (AREA)
- Proteomics, Peptides & Aminoacids (AREA)
- Analytical Chemistry (AREA)
- Zoology (AREA)
- Pathology (AREA)
- Veterinary Medicine (AREA)
- Public Health (AREA)
- Animal Behavior & Ethology (AREA)
- Engineering & Computer Science (AREA)
- Pharmacology & Pharmacy (AREA)
- Medicinal Chemistry (AREA)
- Genetics & Genomics (AREA)
- Immunology (AREA)
- Wood Science & Technology (AREA)
- Biochemistry (AREA)
- Epidemiology (AREA)
- Bioinformatics & Cheminformatics (AREA)
- General Engineering & Computer Science (AREA)
- Hospice & Palliative Care (AREA)
- Microbiology (AREA)
- Biotechnology (AREA)
- Molecular Biology (AREA)
- Biophysics (AREA)
- Physics & Mathematics (AREA)
- Oncology (AREA)
- Chemical Kinetics & Catalysis (AREA)
- General Chemical & Material Sciences (AREA)
- Nuclear Medicine, Radiotherapy & Molecular Imaging (AREA)
- Measuring Or Testing Involving Enzymes Or Micro-Organisms (AREA)
Abstract
The new application that the invention discloses LINC01702 in diagnosing hepatocellular carcinoma.It can detect the expression quantity of LINC01702 in biological sample for the primer and probe of LINC01702 by designing.Judge whether subject suffers from hepatocellular carcinoma by the height of the expression quantity of LINC01702.The studies above achievement according to the present invention, can prepare the product of diagnosing hepatocellular carcinoma, which is suitable for clinically promoting.
Description
Technical field
The invention belongs to fields of biomedicine, are related to LINC01702 answering in the diagnostic kit of preparation hepatocellular carcinoma
With.
Background technique
Hepatocellular carcinoma (hepatocellular carcinoma, HCC) causes to occupy second in dead tumour all.
Cirrhosis is the Major Risk Factors that HCC is formed, incidence 80%-90%, and the principal element of cirrhosis is caused to be B-mode
Hepatites virus infections.The main method for the treatment of HCC includes operation excision, liver transplant, adjuvant treatment and interventional therapy etc. at present.
But HCC can be just diagnosed after related symptoms appearance, lead to 5 years survival rates of HCC patient only 7%.HCC postoperative recurrence and transfer
It is the main reason for causing survival rate low.Although the mechanism of HCC is not yet clear, epithelial-mesenchymal is converted HCC's
It plays a significant role in formation.The traditional serological of HCC diagnoses based on alpha-fetoprotein (crfetoprotein, AFP), but
15%-58% chronic hepatitis B patient and 11%-47% serum of cirrhosis patients AFP level can also increase.Early stage individually surveys
The rate of missed diagnosis for determining AFP is up to 40%, and advanced stage still has 30% patients serum AFP horizontal normal.Though imageological examination has more highly sensitive
Property, but accuracy depends on identifying the ability of neoplastic lesion and nonneoplastic lesion.Result of study shows long-chain non-coding
RNA (long non-coding RNA, 1ncRNA) plays a significant role in HCC generation, while 1ncRNA has high tissue special
The opposite sex is highly expressed in associated tumor tissue, and therefore, 1ncRNA is expected to become the Novel marker of prediction HCC and treatment
Novel targets.
Gene in genome less than 2% is used for coding protein, though most gene, which is transcribed not, has coding
The function of protein, such transcription product are known as non-coding RNA (non-coding RNA, ncRNA).NcRNA is according to length
Difference is divided into short and small ncRNA and LncRNA.LncRNA is a kind of length>200nt, the not molecule of coding protein.LncRNA and
MRNA has more common feature, such as:It is mostly transcribed by rna plymerase ii, by montage, Polyadenylation and 5 ' tailings
?；But LncRNA lacks or seldom has open reading frame, and Transcript Length is short, and positioning in the cell is different from mRNA, more
At nucleus.LncRNA transcriptional level is low compared with encoding egg white gene, and stability is poor, but still has highly conserved region, and this
A little regions are mostly related to the generation of tumour, and 1ncRNA plays an important role in the generation of tumour.
It is different from the position of encoding gene according to LncRNA, it can be divided into just (sense), antisense (antisense), two-way
(bidirectional), 5 kinds of (inYergenic) between (intronic), gene between introne.Adjustment mechanism may relate to：(1)
Neighbouring encoding egg white gene is interfered to express；(2) inhibit polymerase II activity；(3) posttranscriptional modification is participated in；(4) with functional egg
White combination；(5) as the precursor substance of microRNA；(6) it in conjunction with chromosome, is adjusted by signal path.LncRNA is thin
Growth, apoptosis, metabolism, immune response and the tumour of born of the same parents plays important regulative in occurring.Therefore, LncRNA
It is expected to the Novel marker and therapy target as diagnosing tumour.
Summary of the invention
The purpose of the present invention is to provide a kind of long-chain non-coding RNA markers for diagnosing hepatocellular carcinoma.The present invention
Using experiments have shown that expression of the LINC01702 in Tissues of Hepatocellular Carcinoma significantly lower than level in cancer beside organism, because
This can be using LINC01702 as the molecular marker of diagnosing hepatocellular carcinoma.
In order to test above-mentioned purpose, present invention employs following technical solutions：
The present invention provides the reagents of detection long-chain non-coding RNA expression to prepare answering in diagnosis of hepatoma product
With.
Long-chain non-coding RNA of the invention is named as LINC01702 in NCBI.Belong to gene I/D：102724481 compile
The LINC01702 of code product has the transcript of different length, therefore long-chain non-coding RNA-LINC01702 of the invention can be with
Be Genbank accession number XR_947471.2 (with 1641bp length, corresponding DNA sequence dna is as shown in SEQ ID NO.1),
Genbank accession number XR_947473.2 (length with 1573p, corresponding DNA sequence dna is as shown in SEQ ID NO.2),
Genbank accession number XR_426708.3 (with 1080p length, corresponding DNA sequence dna is as shown in SEQ ID NO.3), in
Any one is multiple.
Further, the reagent including the use of SYBR Green, TaqMan probe, molecular beacon, double cross probe or is answered
The PCR amplification primer used when closing probe in detecting LINC01702 expression quantity.
In specific embodiments of the present invention, the primer sequence is as shown in SEQ ID NO.4 and SEQ ID NO.5.
The present invention provides a kind of product for diagnosis of hepatoma, the product includes detection LINC01702 expression
Horizontal reagent.
Further, the reagent includes SYBR Green, TaqMan probe, molecular beacon, double cross probe or compound spy
Needle detects the PCR amplification primer used when LINC01702 expression quantity.
In specific embodiments of the present invention, the primer sequence is as shown in SEQ ID NO.4 and SEQ ID NO.5.
Further, mentioned-above product includes but is not limited to chip, kit, test paper or high-flux sequence platform；It is high
Flux microarray dataset is a kind of tool of special diagnosing hepatocellular carcinoma, with the development of high throughput sequencing technologies, to a people
Rna expression spectrum building will become very easily work.It is composed by the rna expression of comparison Disease and normal population,
The exception for being easy to analyze which RNA is related to disease.Therefore, the unconventionality expression of LINC01702 is known in high-flux sequence
The purposes for also belonging to LINC01702 related to hepatocellular carcinoma, equally within protection scope of the present invention.
The kit include detect LINC01702 expression quantity reagent, the reagent include with LINC01702 or its
The nucleic acid that DNA sequence dna combines, the nucleic acid include SYBR Green, TaqMan probe, molecular beacon, double cross probe or multiple
The PCR amplification primer used when closing probe in detecting LINC01702 expression quantity.
The chip includes the reagent for detecting LINC01702 expression quantity, and the reagent includes and LINC01702 or its DNA
The nucleic acid that sequence combines, the nucleic acid includes the probe for being able to detect LINC01702 expression quantity.
The test paper includes the reagent for detecting LINC01702 expression quantity, and the reagent includes and LINC01702 or its DNA
The nucleic acid that sequence combines, the nucleic acid includes the probe for being able to detect LINC01702 expression quantity.
The present invention provides a kind of for inhibiting the pharmaceutical composition of hepatocellular carcinoma, and described pharmaceutical composition includes
The agonist of LINC01702.
Further, the agonist is unrestricted, as long as can promote LINC01702 expression or promotion
LINC01702 functional activity.
The agonist includes LINC01702 itself, or can express the carrier of LINC01702, promotion property miRNA,
Promoted type transcription regulaton factor, promoted type small molecule compound.
Pharmaceutical composition of the invention can be used as medicine and be administered alone or apply together with other medicines.It can be with this hair
The other medicines that bright pharmaceutical composition is applied together are unrestricted, as long as it does not damage therapeutic or preventative medicine of the invention
The effect of compositions.
Pharmaceutical composition of the invention can be prepared into various dosage forms as needed.Including but not limited to, percutaneous, mucous membrane, nose,
Buccal, the sublingual or oral tablet used, solution, granule, patch, paste, capsule, aerosol or suppository.
The administration method of pharmaceutical composition of the invention is unrestricted, as long as it can play desired therapeutic effect or prevention
Effect, including but not limited to intravenously, in peritonaeum, intraocularly, intra-arterial, intrapulmonary is taken orally, in vesicle, intramuscular, and tracheae
Interior, subcutaneous, local by pleura by skin, sucking, by mucous membrane, skin, stomach is intra-articular, intra-ventricle, directly
Intestines, vagina, in skull, in urethra, in liver, in tumor.In some cases, it can systematically be administered.It is office in some cases
Portion it is administered.
The dosage of pharmaceutical composition of the invention is unrestricted, as long as obtaining desired therapeutic effect or preventive effect i.e.
Can, appropriate determination can be carried out according to symptom, gender, age etc..Therapeutic agent composition of the invention or prophylactic agent combination
The dosage of object, which can be used, for example determines the therapeutic effect of disease or preventive effect as index.
The present invention also provides mentioned-above long-chain non-coding RNA answering in the drug of preparation treatment hepatocellular carcinoma
With.
The present invention also provides application of the mentioned-above agonist in the drug of preparation treatment hepatocellular carcinoma.
The present invention also provides a kind of methods of diagnosing hepatocellular carcinoma, and described method includes following steps：
(1) sample of subject is obtained；
(2) in test sample LINC01702 expression；
(3) by the expression of the LINC01702 measured, whether illness associates with subject；
(4) compared with normal population, the expression of LINC01702 is significantly reduced, then the subject, which is judged, has suffered from
Liver cancer, or the risk with liver cancer is high in the future.
The present invention also provides a kind of suppressing methods of hepatocellular carcinoma, and the method includes promoting LINC01702 expression quantity
Or promote the functional activity of LINC01702.
The present invention also provides a kind of screening techniques of drug for treating hepatocellular carcinoma, can be by thin to hepatocellular carcinoma
The expression that born of the same parents add some period measurement LINC01702 after testing drug improves tumor development to measure tumour medicine
Effect.More specifically, when the expression of LINC01702 when increasing after adding or applying testing drug or restores normal
When horizontal, therapeutic agent of the drug as treatment hepatocellular carcinoma may be selected.
" treatment " used herein is covered treatment-related in such as mankind of the mammal with related disease or illness
Disease or morbid state, and including：
(1) prevent disease or morbid state occurs in mammals, especially when the mammal is susceptible in the disease
Diseased state, but when being not yet diagnosed with this morbid state；
(2) inhibit disease or morbid state, that is, prevent its generation；Or
(3) alleviate disease or morbid state, even if disease or morbid state subside.
Term " treatment " is usually directed to treatment mankind or animal (for example, being applied by animal doctor), wherein can reach certain pre-
The therapeutic effect of phase, for example, inhibiting the development (including reduce development speed, stop development) of illness, improving illness and healing
Illness.It further include the treatment as precautionary measures (such as prevention).To not yet development be illness but have development be the illness endanger
The purposes of the patient of danger, is also included in term " treatment ".
The advantages of the present invention：
The molecular marker for having found a kind of diagnosing hepatocellular carcinoma of the invention, can be thin in liver using the molecular marker
Born of the same parents carcinogenesis early stage can be used as judging, improve the survival rate of patient.
The therapeutic agent of agonist including LINC01702 of the invention can be used as the medicine of new treatment hepatocellular carcinoma
Object.
Detailed description of the invention
Fig. 1 shows the statistics using QPCR detection LINC01702 expression in Tissues of Hepatocellular Carcinoma and cancer beside organism
Figure.
Specific embodiment
The present invention is described in further detail with reference to the accompanying drawings and examples.Following embodiment is merely to illustrate this
It invents rather than limits the scope of the invention.Test method without specific conditions in embodiment, usually according to conventional strip
Part, such as Sambrook et al., molecular cloning：Laboratory manual (New York:Cold Spring HarborLaboratory
Press, 1989) condition described in, or according to the normal condition proposed by manufacturer.
Embodiment 1 screens the non-long-chain coding RNA of differential expression
1, research object
This research is examined by Hospital Ethical Committee and approval, and all patients sign written informed consent before entering group
Book.4 pairs of liver cancer tissues and cancer beside organism's sample are all from the patient of hospital's row Hepatectomy.Experimental subjects is included in standard such as
Shown in lower：
1) hepatocellular carcinoma is turned out to be through pathological examination；
2) receive hepatectomy for the first time, do not receive chemotherapy before, the treatment such as embolism, radio frequency；
3) this research of patient's voluntary participation, and be able to cooperate and complete coherence check；
4 patients with hepatocellular carcinoma are included in experiment altogether, and non-tumour hepatic tissue is cut from hepatectomy by Tissues of Hepatocellular Carcinoma and cancer
The specimens from pri of art.Medical history information is complete, turns out to be hepatocellular carcinoma through hospital pathology department pathologic finding in postoperative.Tissue specimen
It is cut immediately after operation, is placed in transhipment in liquid nitrogen, is stored in -80 DEG C of refrigerators.
2, tissue RNA is always extracted
Referring to mirVanaTMIsolation Kit (Applied Biosystems, Foster City, CA, USA) explanation
Book extracts total tissue RNA.
(1) lysis/binding buffer (the 1ml lysis/binding buffer/0.1g group of 10 times of volumes is added
Knit) homogenizer thoroughly mixes.
(2) the Homogenate additive of 1/10 volume is added, is vortexed and mixes, places 10 minutes on ice.Operation exists
It carries out on ice.
(3) acid-phenol with lysis (disregarding Homogenate additive) same volume is added：
chloroform(300μ1lysis/300μ1acid-phenol：Chloroform), vortex 30-60 seconds, room temperature 10,000g from
It the heart 5 minutes, if split-phase is bad, is then centrifuged again.It takes supernatant to set in a new pipe, remembers volume.
(4) 1.25 times of 100% ethyl alcohol of volume are added, is vortexed and mixes, cross purification column repeatedly, volume is no more than 700 μ 1, room temperature
10,000g is centrifuged 15 seconds.
(5) 350 μ 1wash 1 are added, are centrifuged 5-10 seconds, cleaning purifying is leant on, and room temperature 10,000g is centrifuged 15 seconds, abandons filtering
Liquid.
(6) 10 μ l and Buffer RDD of DNaseI, 70 μ l is added on film (QIAGEN#79254), and 20-30 DEG C is placed 15 points
Clock.
(7) 350 μ 1wash 1 are added, are centrifuged 5-10 seconds, cleaning purifying is leant on, and filtered fluid is abandoned in 10,000g centrifugations 15 seconds.
(8) 500 μ 1wash 2/3 are added, are centrifuged 5-10 seconds, cleaning purification column is secondary, and filtering is abandoned in 10,000g centrifugations 15 seconds
Liquid is centrifuged 1 minute.
(9) centrifugal column is placed into new collecting pipe, the Elution of 1 95 DEG C of 100 μ preheatings is added in column center
Solution or nuclease-free water, room temperature maximum speed are centrifuged 20-30 seconds, and liquid is the Total extracted in collecting pipe
RNA is placed on -70 DEG C of preservations.
(10) detection of RNA：1 μ 1RNA sample is taken to utilize 2000 spectrophotometer (Thermo of NanoDrop
Scientific, USA) measure concentration and OD260/OD280, the integrality of agarose gel electrophoresis detection RNA.
3, chip detects
The detection of lncRNA+mRNA chip of expression spectrum is starting with the total serum IgE of sample to be tested, carries out amplification in vitro and fluorescence
Label, labeling process use biochip common tags kit.Mainly include the following steps：
(1) reverse transcription synthesizes the first chain cDNA：With Total RNA starting, the T7Oligo (dT) containing T7 promoter sequence
Primer and T7- random primer, both can with the mRNA of junction belt poly (A), can also in conjunction in addition to rRNA it is other without
The RNA of poly (A) synthesizes the first chain cDNA using the first chain Enzyme Mix.
(2) the second chain DNA is synthesized：Second is converted by the RNA chain in DNA-RNA heterozygote with the second chain Enzyme Mix
Chain cDNA, synthetic dsdna.
(3) synthesis cRNA is transcribed in vitro：Using the second chain cDNA as template, cRNA is synthesized using T7Enzyme Mix.
(4) cRNA is purified：Purify column purification cRNA using RNA, except reagents such as salt, enzymes in dereaction, and to cRNA into
Capable quantitative, Quality Control.
(5) reverse transcription：Using cRNA as template, Random Primer is primer, and II enzyme of CbcScript carries out reverse transcription.It is pure
Change the cDNA that recycling reverse transcription obtains and quantifies.
(6) it marks：Using the cDNA product of reverse transcription as template, Random Primer is primer, uses Klenow
Fragment enzymatic synthesis cDNA complementary strand simultaneously mixes the dNTP (Cy3-dCTP, Cy5-dCTP) for having fluorophor, purifies and fixed
Measure marked product.It can be used to chip hybridization with fluorophor DNA.
(7) chip hybridization and cleaning：1 μ L cDNA is taken to be dissolved in hybridization solution, 45 DEG C of hybridized overnights.Chip is taken out in rich Austria
SlideWasher8 chip is washed dry instrument and is cleaned, and cleaning procedure is as follows：Washing lotion I：0.2%SDS, 2 × SSC, 42 DEG C of 120s are clear
It washes 2 times.Washing lotion II：0.2%SDS, 2 × SSC, 42 DEG C of 80s are cleaned 3 times.After the completion of cleaning procedure, centrifuge dripping is to be scanned.
(8) chip scanning, data analysis, differential gene screening：Using Agilent chip scanner (G2565CA) to clear
Chip after washing is scanned, and obtains hybridization picture.Using Agilent Feature Extraction (v10.7) software to miscellaneous
Intersection graph piece is analyzed and extracts data.Then using Agilent GeneSpring software data are normalized and difference
Analysis, and differential gene screening is carried out with GeneSpring GX software.
4, result
Screening criteria：P<0.01, abs (log2 (fold change))>2,311 differential expression lncRNA are filtered out altogether,
Wherein 204 up-regulations, 107 downwards.
2 large sample of embodiment verifies the differential expression LncRNA filtered out
LINC01702 is selected to be verified according to the size of P value based on the selection result of embodiment 1.
1, sample collection
Tissues of Hepatocellular Carcinoma 36 are collected according to the method for embodiment 1, corresponding cancer beside organism 36.
2, it is verified on transcriptional level
2.1 extract tissue RNA
Step is the same as embodiment 1.
2.2 design of primers
According to LINC01702 transcript sequence, by design of primers tool (Primer BLAST) design primer of NCBI,
The primer is the universal primer of tri- kinds of transcripts of LINC01702, and primer sequence is as follows：Upstream primer：5'-
GGCAATGGAGAATATAAAG-3'(SEQ ID NO.4)；Downstream primer：5'-TTCATCTTGTCACTACTT-3'(SEQ ID
NO.5)。
According to SnU6 primers, upstream primer：5'-CTCGCTTCGGCAGCACATATACT-3'(SEQ ID
NO.6)；5'-ACGCTTCACGAATTTGCGTGTC-3'(SEQ ID NO.7).
2.3cDNA synthesis
With the total serum IgE (1 μ g) of extraction for template, following reaction system is added, specially：Buffer
4 μ L,1 μ L, Oligo dT Primer (50 μM) of RT Enzyme Mix, 1 μ L, Random 6mers (100 μM)
1 μ L, with the ddH of no RNA enzyme2O supplies reaction volume for 20 μ L.Above-mentioned mixed liquor is placed in 37 DEG C of 15min, 85 DEG C of 5s, i.e.,
Obtain cDNA.The cDNA can be used for lncRNA Real-time PCR detection.
2.4Real-time PCR
It is glimmering using Beijing Kang Run Cheng Ye Biotechnology Co., Ltd 2 × RealStar Power SYBR Mixture UNG
Light quantification kit (article No. A312) carries out fluorescent quantitative PCR by template of the cDNA of reverse transcription.
Quantitative fluorescent PCR system is as shown in table 1.
1 quantitative fluorescent PCR system of table
| Reagent | Dosage |
| 2*MIX | 10μL |
| 50*ROX | 2μL |
| Primer | Each 1 μ L |
| DNA profiling | 1μL |
| RNase Free water | Polishing is to 20 μ L |
Quantitative fluorescent PCR program：95 DEG C of 30s initial denaturations connect 40 circulations：95 DEG C of 30s, 60 DEG C of 30s.
According to the relative quantification formula of qRT-PCR：2- △ Ct calculates separately out LINC01702 in patients with hepatocellular carcinoma cancer
Expression in tissue and cancer beside organism.The display of this experimental result：LINC01702 compares cancer in 36 Tissues of Hepatocellular Carcinoma
Side is organized, and is lowered in 32, positive rate, and=downward expression number of cases/always detects number of cases × 100%=32/36=88.9%.
Statistical result such as Fig. 1 is shown, compared with Tissues of Hepatocellular Carcinoma, LINC01702 expression is significantly reduced in cancer beside organism, difference
With statistical significance (P<0.05).These results suggest that LINC01702 common lower expression in Tissues of Hepatocellular Carcinoma.
LINC01702 can be used as the new tumor markers of diagnosis of hepatoma, the screening for hepatocellular carcinoma.
The explanation of above-described embodiment is only intended to understand method and its core concept of the invention.It should be pointed out that for this
For the those of ordinary skill in field, without departing from the principle of the present invention, several improvement can also be carried out to the present invention
And modification, these improvement and modification will also be fallen into the protection scope of the claims in the present invention.
Sequence table
<110>Taishan Hospital, Hospital Attached to Taishan Medical College
<120>Application of the LINC01702 in the diagnostic kit of preparation hepatocellular carcinoma
<160> 7
<170> SIPOSequenceListing 1.0
<210> 1
<211> 1641
<212> DNA
<213>Source of people (Homo sapiens)
<400> 1
tacaggcata cgccaccgca cctggcccaa attattaatt gtacgtattc tcattgttct 60
gcttcttcca agaaaactga agtcacggta ttctgaagac tagagatgat tcaacaggct 120
gtgaatctat tccatttgga atcctactgg gcctgatctg ttttccattg ccaaggcact 180
gctgtttaag ctataccata tgaagcaccc tccctaaagg ctcaggggcc atagtgaaag 240
aggacacatg ggattgtaag agccagatta tgggggtgga gggtaaaggc tgaagcaagt 300
ccatcttgga tgctaactca ctatgttagc ttctgattaa ccccagttcc aggaaggctg 360
ctgaaatttc cagtttatgt gttgttcctt gtgtaagagc atggacttac tgcaaatcct 420
gcccttaggg caaattccta cctattcctg cggaagcatg tgtatcattc ccctatggtt 480
tataacccct gagtctgggg gtaatggcac ggggacccac cagcttgtct gccgccatct 540
ggggtacagt gctggaagcg gggatacagg gacaaataag acacagatcc tgttcctaaa 600
gagagtcttt tctaaggcac caactacttc cttcaagcag cagccagaga tctttgctta 660
atggctgaga aacacagagg agagacccag agagaaggag aaggccgcat gaagatggag 720
cagagactgg agttattcag ccaagtcaag gaatgcctgg agcaccagaa gccggaagag 780
gcaaggaagg atcctccctt agagcatttg gaagagtgtg gccctgccaa caccttgact 840
tcaaacttct ggcctccata actatgaaat aaattcttgt tgttttaagc caccaagttt 900
gtggtaactt cttacagcaa ccctaggaaa ttaacacaca gagtgtcagt tacctaaaca 960
ttgtgagcct cagttacctt atctgaaaat ggggctaaaa cttagaccta ccttttaggg 1020
ctgctttaag gagtaataga aattatacat gggaattaca tggcaccaag ttgttcctta 1080
atagagggaa actgttatga tggcggaaga gcagtaatga aatactgaat tcatccagca 1140
gatggcagaa gatggatttg caaacaagag gacttcataa ggtttgctgc ccagcaatgc 1200
agctccaggc ctaaagtagc cttcctagct cacagactga tgaggattaa ccctcgccaa 1260
ctcaagtgtg ctctcagctc tgtagaaact aaactaattc tgttttcctt tgaccattgt 1320
acaccacaga ggctggcaat ggagaatata aagagtggtg caacagaaaa tacctgggtt 1380
ctaggctcag ctctgtcacc tcattaggat tcagtttgcc catctgtaaa gtagtgacaa 1440
gatgaaatac caagtgtaac gtaattaaag ggctgttgtg ataccaaaat aacataatgt 1500
gtaaaagtgc taaaaacaag atgccataaa gcacagaagg ggcccaggag agattaattc 1560
atggagaact tactcgtttt tttgagacag agtctcactg tgttgctcag gctgcagtgc 1620
agtggcgcga tctcagctaa c 1641
<210> 2
<211> 1573
<212> DNA
<213>Source of people (Homo sapiens)
<400> 2
tacgccaccg cacctggccc aaattattaa ttgtacgtat tctcattgtt ctgcttcttc 60
caagaaaact gaagtcacgg tattctgaag actagagatg attcaacagg ctgtgaatct 120
attccatttg gaatcctact gggcctgatc tgttttccat tgccaaggca ctgctgttta 180
agctatacca tatgaagcac cctccctaaa ggctcagggg ccatagtgaa agaggacaca 240
tgggattgta agagccagat tatgggggtg gagggtaaag gctgaagcaa gtccatcttg 300
gatgctaact cactatgtta gcttctgatt aaccccagtt ccaggaaggc tgctgaaatt 360
tccagtttat gtgttgttcc ttgtgtaaga gcatggactt actgcaaatc ctgcccttag 420
ggcaaattcc tacctattcc tgcggaagca tgtgtatcat tcccctatgg tttataaccc 480
ctgagtctgg gggtaatggc acggggaccc accagcttgt ctgccgccat ctgggagtct 540
tttctaaggc accaactact tccttcaagc agcagccaga gatctttgct taatggctga 600
gaaacacaga ggagagaccc agagagaagg agaaggccgc atgaagatgg agcagagact 660
ggagttattc agccaagtca aggaatgcct ggagcaccag aagccggaag aggcaaggaa 720
ggatcctccc ttagagcatt tggaagagtg tggccctgcc aacaccttga cttcaaactt 780
ctggcctcca taactatgaa ataaattctt gttgttttaa gccaccaagt ttgtggtaac 840
ttcttacagc aaccctagga aattaacaca cagagtgtca gttacctaaa cattgtgagc 900
ctcagttacc ttatctgaaa atggggctaa aacttagacc taccttttag ggctgcttta 960
aggagtaata gaaattatac atgggaatta catggcacca agttgttcct taatagaggg 1020
aaactgttat gatggcggaa gagcagtaat gaaatactga attcatccag cagatggcag 1080
aagatggatt tgcaaacaag aggacttcat aaggtttgct gcccagcaat gcagctccag 1140
gcctaaagta gccttcctag ctcacagact gatgaggatt aaccctcgcc aactcaagtg 1200
tgctctcagc tctgtagaaa ctaaactaat tctgttttcc tttgaccatt gtacaccaca 1260
gaggctggca atggagaata taaagagtgg tgcaacagaa aatacctggg ttctaggctc 1320
agctctgtca cctcattagg attcagtttg cccatctgta aagtagtgac aagatgaaat 1380
accaagtgta acgtaattaa agggctgttg tgataccaaa ataacataat gtgtaaaagt 1440
gctaaaaaca agatgccata aagcacagaa ggggcccagg agagattaat tcatggagaa 1500
cttactcgtt tttttgagac agagtctcac tgtgttgctc aggctgcagt gcagtggcgc 1560
gatctcagct aac 1573
<210> 3
<211> 1080
<212> DNA
<213>Source of people (Homo sapiens)
<400> 3
agcaattctc ctgtctcagc ctcctgagta gctggattac agagtctttt ctaaggcacc 60
aactacttcc ttcaagcagc agccagagat ctttgcttaa tggctgagaa acacagagga 120
gagacccaga gagaaggaga aggccgcatg aagatggagc agagactgga gttattcagc 180
caagtcaagg aatgcctgga gcaccagaag ccggaagagg caaggaagga tcctccctta 240
gagcatttgg aagagtgtgg ccctgccaac accttgactt caaacttctg gcctccataa 300
ctatgaaata aattcttgtt gttttaagcc accaagtttg tggtaacttc ttacagcaac 360
cctaggaaat taacacacag agtgtcagtt acctaaacat tgtgagcctc agttacctta 420
tctgaaaatg gggctaaaac ttagacctac cttttagggc tgctttaagg agtaatagaa 480
attatacatg ggaattacat ggcaccaagt tgttccttaa tagagggaaa ctgttatgat 540
ggcggaagag cagtaatgaa atactgaatt catccagcag atggcagaag atggatttgc 600
aaacaagagg acttcataag gtttgctgcc cagcaatgca gctccaggcc taaagtagcc 660
ttcctagctc acagactgat gaggattaac cctcgccaac tcaagtgtgc tctcagctct 720
gtagaaacta aactaattct gttttccttt gaccattgta caccacagag gctggcaatg 780
gagaatataa agagtggtgc aacagaaaat acctgggttc taggctcagc tctgtcacct 840
cattaggatt cagtttgccc atctgtaaag tagtgacaag atgaaatacc aagtgtaacg 900
taattaaagg gctgttgtga taccaaaata acataatgtg taaaagtgct aaaaacaaga 960
tgccataaag cacagaaggg gcccaggaga gattaattca tggagaactt actcgttttt 1020
ttgagacaga gtctcactgt gttgctcagg ctgcagtgca gtggcgcgat ctcagctaac 1080
<210> 4
<211> 19
<212> DNA
<213>Artificial sequence (Artificial Sequence)
<400> 4
ggcaatggag aatataaag 19
<210> 5
<211> 18
<212> DNA
<213>Artificial sequence (Artificial Sequence)
<400> 5
ttcatcttgt cactactt 18
<210> 6
<211> 23
<212> DNA
<213>Artificial sequence (Artificial Sequence)
<400> 6
ctcgcttcgg cagcacatat act 23
<210> 7
<211> 22
<212> DNA
<213>Artificial sequence (Artificial Sequence)
<400> 7
acgcttcacg aatttgcgtg tc 22
Claims (10)
1. the reagent of detection long-chain non-coding RNA expression is preparing the application in diagnosis of hepatoma product；The non-volume of long-chain
Code RNA is LINC01702, the encoding gene Gene ID of the long-chain non-coding RNA：105373113.
2. application according to claim 1, which is characterized in that the reagent is visited including the use of SYBR Green, TaqMan
Needle, molecular beacon, double cross probe or combined probe detect the pcr amplification primer used when the long-chain non-coding RNA expression quantity
Object.
3. application according to claim 2, which is characterized in that the primer sequence such as SEQ ID NO.4 and SEQ ID
Shown in NO.5.
4. application according to any one of claim 1-3, which is characterized in that the product include kit, chip or
Test paper.
5. a kind of product for diagnosis of hepatoma, which is characterized in that the product includes any in detection claim 1-4
The reagent of long-chain non-coding RNA expression described in.
6. product according to claim 5, which is characterized in that the reagent includes SYBR Green, TaqMan probe, divides
Sub- beacon, double cross probe or combined probe detect the PCR amplification primer used when the long-chain non-coding RNA expression quantity.
7. product according to claim 6, which is characterized in that the primer sequence such as SEQ ID NO.4 and SEQ ID
Shown in NO.5.
8. a kind of for inhibiting the pharmaceutical composition of hepatocellular carcinoma, which is characterized in that described pharmaceutical composition includes claim
The agonist of long-chain non-coding RNA described in any one of 1-4.
9. pharmaceutical composition according to claim 8, which is characterized in that the agonist includes promoting long-chain non-coding
The reagent of rna level, or promote the reagent of the long-chain non-coding RNA functional activity.
10. application of the long-chain non-coding RNA described in claim 1 in the drug of preparation treatment hepatocellular carcinoma.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201810833682.2A CN108841961B (en) | 2018-07-26 | 2018-07-26 | Application of the LINC01702 in the diagnostic kit of preparation hepatocellular carcinoma |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201810833682.2A CN108841961B (en) | 2018-07-26 | 2018-07-26 | Application of the LINC01702 in the diagnostic kit of preparation hepatocellular carcinoma |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN108841961A true CN108841961A (en) | 2018-11-20 |
| CN108841961B CN108841961B (en) | 2019-07-05 |
Family
ID=64195737
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN201810833682.2A Expired - Fee Related CN108841961B (en) | 2018-07-26 | 2018-07-26 | Application of the LINC01702 in the diagnostic kit of preparation hepatocellular carcinoma |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN108841961B (en) |
Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN112553343A (en) * | 2021-01-05 | 2021-03-26 | 武汉大学 | Blood molecular marker for hepatocellular carcinoma auxiliary diagnosis, primer and application thereof |
Citations (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN106995858A (en) * | 2017-06-01 | 2017-08-01 | 北京泱深生物信息技术有限公司 | A kind of lncRNA related to liver cancer diagnosis and treatment |
| CN107083433A (en) * | 2017-06-01 | 2017-08-22 | 北京泱深生物信息技术有限公司 | Applications of the lncRNA in liver cancer diagnosis and treatment |
| CN107267625A (en) * | 2017-07-06 | 2017-10-20 | 王冬国 | Purposes of the lncRNA as biomarker in liver cancer diagnosis and treatment |
| CN107586781A (en) * | 2017-10-25 | 2018-01-16 | 中国人民解放军第三〇二医院 | Liver cancer marker lncRNA ENST00000620463.1 and its application |
| CN107858434A (en) * | 2017-12-22 | 2018-03-30 | 复旦大学附属肿瘤医院 | Applications of the lncRNA in diagnosing cancer of liver and prognosis prediction |
-
2018
- 2018-07-26 CN CN201810833682.2A patent/CN108841961B/en not_active Expired - Fee Related
Patent Citations (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN106995858A (en) * | 2017-06-01 | 2017-08-01 | 北京泱深生物信息技术有限公司 | A kind of lncRNA related to liver cancer diagnosis and treatment |
| CN107083433A (en) * | 2017-06-01 | 2017-08-22 | 北京泱深生物信息技术有限公司 | Applications of the lncRNA in liver cancer diagnosis and treatment |
| CN107267625A (en) * | 2017-07-06 | 2017-10-20 | 王冬国 | Purposes of the lncRNA as biomarker in liver cancer diagnosis and treatment |
| CN107586781A (en) * | 2017-10-25 | 2018-01-16 | 中国人民解放军第三〇二医院 | Liver cancer marker lncRNA ENST00000620463.1 and its application |
| CN107858434A (en) * | 2017-12-22 | 2018-03-30 | 复旦大学附属肿瘤医院 | Applications of the lncRNA in diagnosing cancer of liver and prognosis prediction |
Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN112553343A (en) * | 2021-01-05 | 2021-03-26 | 武汉大学 | Blood molecular marker for hepatocellular carcinoma auxiliary diagnosis, primer and application thereof |
Also Published As
| Publication number | Publication date |
|---|---|
| CN108841961B (en) | 2019-07-05 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| AU2013281355B2 (en) | Targeted RNA-seq methods and materials for the diagnosis of prostate cancer | |
| CN117965741A (en) | Kit or device for detecting pancreatic cancer and detection method | |
| Elbere et al. | Significantly altered peripheral blood cell DNA methylation profile as a result of immediate effect of metformin use in healthy individuals | |
| CN109055555B (en) | Lung cancer early stage metastasis diagnosis marker and kit and application thereof | |
| Shimizu et al. | Identification of intragenic methylation in the TUSC1 gene as a novel prognostic marker of hepatocellular carcinoma | |
| CN111269985B (en) | Application of hsa _ circRNA6448-14 in diagnosis and prognosis prediction of esophageal squamous cell carcinoma | |
| CN108949969B (en) | Application of long-chain non-coding RNA in colorectal cancer | |
| WO2013148151A1 (en) | Plasma microribonucleic acids as biomarkers for endometriosis and endometriosis-associated ovarian cancer | |
| CN106834470A (en) | Purposes of the miRNA in cancer diagnosing kit is prepared | |
| CN107312865B (en) | Purposes of the LOC100130111 in preparation osteosarcoma diagnostic products, therapeutic agent | |
| Kupferman et al. | Molecular analysis of anoikis resistance in oral cavity squamous cell carcinoma | |
| CN108624693B (en) | MiR-577 is preparing the application in diagnosis of nephropathy marker | |
| CN107475386B (en) | Long-chain non-coding RNA marker for diagnosis and treatment osteosarcoma | |
| CN108841961B (en) | Application of the LINC01702 in the diagnostic kit of preparation hepatocellular carcinoma | |
| CN105154541A (en) | Application of miRNA to diagnosis and treatment of acute myelogenous leukemia | |
| CN105886625B (en) | Application of the CHKA gene in preparation cancer of the esophagus diagnosis and treatment product | |
| CN111979315A (en) | Application of annular TP63 as lung squamous carcinoma diagnosis or treatment target | |
| CN107365859B (en) | Molecular markers of the LncRNA as diagnosis and treatment osteosarcoma | |
| JP4967112B2 (en) | Tumor markers for head and neck cancer | |
| US20120058477A1 (en) | Method for detecting the expression of cyclin b2 gene by real-time quantitative pcr | |
| WO2016179814A1 (en) | Gene related to papillary thyroid cancer | |
| CN104450887A (en) | DNA probe and gene chip for detecting endometrial carcinoma and application thereof | |
| JP5897823B2 (en) | Bladder cancer diagnostic composition and method | |
| CN108118093A (en) | Purposes of the LINC00426 in bone and flesh tumor metastasis diagnostic products are prepared | |
| WO2006091049A1 (en) | Protein associated with colorectal cancer, polynucleotide including single-nucleotide polymorphism associated with colorectal cancer, microarray and diagnostic kit including the same, and method of diagnosing colorectal cancer using the same |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| PB01 | Publication | ||
| PB01 | Publication | ||
| SE01 | Entry into force of request for substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| GR01 | Patent grant | ||
| GR01 | Patent grant | ||
| CF01 | Termination of patent right due to non-payment of annual fee |
Granted publication date: 20190705 Termination date: 20200726 |
|
| CF01 | Termination of patent right due to non-payment of annual fee |