CN105154540B - The diagnosis and treatment target of PAEP gene and its expression product as osteoarthritis - Google Patents
The diagnosis and treatment target of PAEP gene and its expression product as osteoarthritis Download PDFInfo
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Abstract
The invention discloses a kind of molecular marker-PAEP genes for osteoarthritis early diagnosis.Experiments have shown that; the mRNA level in-site of PAEP gene in Human Osteoarthritis synovial tissue is substantially less than normal synovial tissue; therefore it may determine that subject whether there is the risk with osteoarthritis by the expression of PAEP gene in measurement subject synovial tissue, or may determine that whether subject has suffered from osteoarthritis.PAEP gene, which can be used for preparing to be applied to Human Osteoarthritis or be applied to, suffers from the high crowd of osteoarthritis risk, for treating osteoarthritis or preventing the generation of osteoarthritis.The present invention provides new diagnostic method for clinically diagnosis osteoarthritis and provides new drug candidate for treatment osteoarthritis.
Description
Technical field
The present invention relates to field of biotechnology, more particularly to people PAEP gene in the diagnosis, treatment of osteoarthritis
Purposes.
Background technique
Osteoarthritis (Osterarthritis, OA) is one group
The clinical syndrome of major pathologic features is a kind of common disease, is a kind of chronic, progressivity joint after mostly occurring in the middle age
Lesion.Occur mainly in articular cartilage.Its show be mainly articular cartilage destroyed, cartilage surface lose homogenieity, interruption,
Patch recess, fracture and ulcer.Recently some research shows that the generation of osteoarthritis is related with inherent cause.Synovial membrane is for maintaining
The normal function in joint, and play an important role during the occurrence and development of osteoarthritis disorders.
The inspection of iconography is clinically based primarily upon for the diagnosis of osteoarthritis at present.Between x-ray performance predominantly joint
Gap is narrow, and subchondral bony sclerosis and cystis degeneration, joint margins spur are formed, and articular surface withers, deforms and subluxation of joint etc..
MRI can show the lesion of the joint structures such as early stage cartilage, meniscus, be conducive to early diagnose.CT is used for the diagnosis of discopathy, excellent
In x-ray.Joint space narrows, Subchondral bone sclerosis, and marginality spur spinal joint is linked to be bone bridge.Also visible bone cyst and abnormal
Shape.As found, these variations can be used as the foundation of diagnosis and the degree of estimation joint injury.OA is generally no or seldom in early stage
There is symptom, only in the activity of inflammation secondary, pain or influence joint, patient just looks for a doctor.At this point, joint injury occurs
Long.Because method of the exploitation for the diagnosis of early stage osteoarthritis is a problem to be solved.
Summary of the invention
In order to make up for the deficiencies of the prior art, the purpose of the present invention is to provide one kind can be used for osteoarthritis
The molecular marker of (Osterarthritis, OA) early diagnosis.Compared to the diagnostic method of traditional osteoarthritis, gene is used
Marker come diagnose osteoarthritis have timeliness, specificity and sensitivity, thus make patient disease early stage can know
Disease risks take corresponding prevention and treatment measure for risk height.
To achieve the goals above, the present invention adopts the following technical scheme:
The answering in the product of preparation diagnosis osteoarthritis the present invention provides a kind of people PAEP gene and its expression product
With.
Further, diagnostic products mentioned above include: by RT-PCR, real-time quantitative PCR, immune detection, original position
Hybridization or chip detect the expression of PAEP gene and its expression product to diagnose the product of osteoarthritis.
Further, the product with RT-PCR diagnosis osteoarthritis includes at least drawing for a pair of of specific amplified PAEP gene
Object；The product with real-time quantitative PCR diagnosis osteoarthritis includes at least the primer of a pair of of specific amplified PAEP gene；It is described
Product with immune detection diagnosis osteoarthritis includes: the antibody in conjunction with PAEP protein-specific；It is described to be examined in situ hybridization
The arthritic product of knochenbruch includes: the probe with the nucleic acid array hybridizing of PAEP gene；It is described to diagnose osteoarthritis with chip
Product includes: protein chip and genetic chip；Wherein, protein chip includes the antibody in conjunction with PAEP protein-specific, gene
Chip includes the probe with the nucleic acid array hybridizing of PAEP gene.
Preferably, the product includes chip, kit.
The present invention also provides application of the people PAEP gene in high-flux sequence platform.With high throughput sequencing technologies
Development will become very easily work to the building of the gene expression profile of a people.Pass through comparison Disease and normal person
The gene expression profile of group, the exception for being easy to analyze which gene are related to disease.Therefore, people is known in high-flux sequence
The exception of the PAEP gene purposes for also belonging to people's PAEP gene related to osteoarthritis, equally protection scope of the present invention it
It is interior.
The present invention also provides the application of people PAEP gene and its expression product in the drug of preparation treatment osteoarthritis.
Further, the drug includes: that the drug of the gene of PAEP containing someone, the carrier of carrier's PAEP gene or host are thin
Born of the same parents be formed by drug, people PAEP pharmaceutical grade protein or other can promote the drug of PAEP gene expression.Drug of the invention
The missing or deficiency that can be used for supplementing endogenic people PAEP albumen, by improving the expression of people PAEP albumen, to treat
Because of osteoarthritis caused by people's PAEP hypoproteinosis.
The carrier of the present invention for carrying gene is various carriers known in the art, such as commercially available carrier including plasmid,
Clay, bacteriophage, virus etc..
In the present invention, term " host cell " includes prokaryotic cell and eukaryocyte.Common prokaryotic host cell
Example includes Escherichia coli, hay bacillus etc..Common eukaryotic host cell includes yeast cells, insect cell and mammal
Cell.Preferably, the host cell is eukaryocyte, such as Chinese hamster ovary celI, COS cell.
Further, drug of the invention further includes pharmaceutically acceptable carrier, carrier, and this kind of carrier includes (but not
It is limited to): diluent, excipient such as water etc., filler such as starch, sucrose etc.；Adhesive such as cellulose derivative, alginates, bright
Glue and polyvinylpyrrolidone；Wetting agent such as glycerol；Disintegrating agent such as agar, calcium carbonate and sodium bicarbonate；Sorbefacient quaternary ammonium
Compound；Surfactant such as hexadecanol；Absorption carrier such as kaolin and soap clay；Lubricant such as talcum powder, calcium stearate
With magnesium, polyethylene glycol etc..
Drug of the invention, which imports tissue or the mode of cell, can be divided into external or intracorporal mode.Vitro formats
Drug including the drug or the protein of PAEP containing someone that will contain someone's PAEP gene imports in cell, then by cell transplantation or
It feeds back in vivo.Internal mode includes directly infusing the drug of the drug of the gene of PAEP containing someone or the protein of PAEP containing someone
Enter in in-vivo tissue.
Drug of the invention can also be with the drug combination of other treatment osteoarthritis, and a variety of Drug combinations can be significantly
Mention the success rate for the treatment of.
The present invention also provides a kind of chips for diagnosing osteoarthritis, and the chip includes genetic chip, protein-chip；
The genetic chip includes solid phase carrier and the oligonucleotide probe for being fixed on solid phase carrier, and the oligonucleotide probe includes
For detecting the oligonucleotide probe for PAEP gene of PAEP gene transcription level；The protein-chip includes that solid phase carries
Body and be fixed on solid phase carrier PAEP albumen specific antibody.
Further, the genetic chip can be used for detecting multiple genes including people's PAEP gene (for example, closing with bone
The scorching relevant multiple genes of section) expression.The protein-chip can be used for detecting more including people's PAEP albumen
The expression of a protein (such as multiple protein relevant to osteoarthritis).By by multiple with osteoarthritis mark
Object detects simultaneously, is greatly improved the accuracy rate of osteoarthritis diagnosis.
The present invention provides a kind of kits for diagnosing osteoarthritis, and the kit includes gene detecting kit and egg
White immunity detection reagent；The gene detecting kit includes the reagent for detecting PAEP gene transcription level；The egg
White immunity detection reagent includes the specific antibody of PAEP albumen.
Further, the reagent includes using RT-PCR, real-time quantitative PCR, immune detection, in situ hybridization or chip method
Detect reagent needed for PAEP gene expression dose process.Preference, the reagent include for PAEP gene primer and/
Or probe.Be easy to design according to the nucleotide sequence of PAEP gene the primer that can be used for detecting PAEP gene expression dose and
Probe.
It can be DNA, RNA, DNA-RNA chimera with the probe of the nucleic acid array hybridizing of PAEP gene, PNA or other spreads out
Biology.There is no limit appoint as long as completing specific hybrid, specifically binding with purpose nucleotide sequence the length of the probe
What length is ok.The length of the probe can be as short as 25,20,15,13 or 10 bases longs.Equally, the length of the probe
Degree can be grown to 60,80,100,150,300 base-pairs or longer or even whole genes.Since different probe lengths is to hybridization
Efficiency, signal specificity have different influences, and the length of the probe is typically at least 14 base-pairs, and longest is usually no more than
30 base-pairs, the length complementary with purpose nucleotide sequence are best with 15-25 base-pair.The probe self-complementary sequences
Most preferably less than 4 base-pairs, in order to avoid influence hybridization efficiency.
Further, the specific antibody of the PAEP albumen includes monoclonal antibody, polyclonal antibody.The PAEP albumen
Specific antibody include complete antibody molecule, antibody any segment or modification (for example, chimeric antibody, scFv, Fab, F
(ab ') 2, Fv etc..As long as the segment can retain the binding ability with PAEP albumen.Antibody for protein level
Preparation when well known to a person skilled in the art and the present invention may use any method to prepare the antibody.
In the context of the present invention, " PAEP gene " includes people PAEP gene and any function of people's PAEP gene etc.
The polynucleotides of jljl.PAEP gene includes and PAEP gene (NC_ in the public GenBank GeneBank in the current world
000009.12) DNA sequence dna has 70% or more homology, and encodes the DNA sequence dna of identical function protein；
Preferably, the coded sequence of PAEP gene includes following any DNA molecular:
(1) DNA sequence dna shown in SEQ ID NO.1 in sequence table；
(2) under strict conditions with 1) DNA sequence dna that limits hybridizes and encodes the DNA sequence dna of identical function protein；
(3) DNA sequence dna limited with (1) or (2) has 70%, preferably, 90% or more homology, and encodes identical function
The DNA molecular of energy protein.
In specific embodiments of the present invention, the coded sequence of the PAEP gene is DNA shown in SEQ ID NO.1
Sequence.
In the context of the present invention, PAEP gene expression product includes the part of people PAEP albumen and people's PAEP albumen
Peptide.The partial peptide of the PAEP albumen contains functional domain relevant to osteoarthritis.
" PAEP albumen " includes any functional equivalent of people PAEP albumen and people's PAEP albumen.The functional equivalent
Including people PAEP albumen conservative variation protein or its active fragment or its reactive derivative, allelic variant is naturally dashed forward
Variant, induced mutants, can be with the encoded protein of DNA of the DNA hybridization of people PAEP under high or low stringent condition.
Preferably, PAEP albumen is the protein with following amino acid sequences:
(1) protein that the amino acid sequence shown in SEQ ID NO.2 in sequence table forms；
(2) amino acid sequence shown in SEQ ID NO.2 by the substitution of one or several amino acid residues and/or is lacked
Lose and/or addition and with the ammonia with the same function as shown in SEQ ID NO.2 of amino acid sequence shown in SEQ ID NO.2
Protein derived from base acid sequence.The number for the amino acid for replacing, lacking or adding is usually 1-50, preferably 1-30
It is a, more preferably 1-20, most preferably 1-10.
(3) there is at least 80% homology (also known as sequence identity) with amino acid sequence shown in SEQ ID NO.2,
It is highly preferred that the homology with amino acid sequence at least about 90% to 95% shown in SEQ ID NO.2, Chang Wei 96%, 97%,
98%, the polypeptide that the amino acid sequence of 99% homology is constituted.
In specific embodiments of the present invention, the PAEP albumen is with amino acid sequence shown in SEQ ID NO.2
The protein of column.
It is known that, conventionally, the modification of one or more amino acid will not influence the function of protein in a protein.
Those skilled in the art can approve the amino acid for changing single amino acids or small percentage or to amino acid sequence it is individual add,
Missing, insertion, replacement are conservative modifications, and wherein the change of protein generates the protein with identity function.Function phase is provided
As the Conservative substitution tables of amino acid be well known in the art.
Example by one amino acid of addition or the protein of more amino acid modification is the fusion of PAEP albumen
Albumen.For the peptide or protein with PAEP protein fusion, there is no limit as long as resulting fusion protein retains PAEP albumen
Biological activity.
PAEP albumen of the invention also includes the non-conservative modification to amino acid sequence shown in SEQ ID NO.2, as long as
Protein by modification still is able to retain the biological activity of PAEP albumen.It is mutated in such modification protein
Amino acid number is usually 10 perhaps less such as 6 perhaps less such as 3 or less.
In the context of the present invention, " diagnosis osteoarthritis " both includes judging whether subject has suffered from Bones and joints
Inflammation also includes the risk that judges subject and whether there is with osteoarthritis.
In the context of the present invention, " treatment osteoarthritis " divides from the state change of disease, may include disease
Alleviate, the complete healing of disease.
The advantages of the present invention:
Present invention firstly discovers that PAEP gene expression is related to osteoarthritis, by detection subject synovial tissue
The expression of PAEP, it can be determined that whether subject suffers from osteoarthritis or judge that subject whether there is with osteoarthritis
Risk, so that clinician be instructed to provide prevention scheme or therapeutic scheme to subject.
Present invention finds a kind of new molecular marked compound-PAEP genes, and compared to traditional detection means, gene diagnosis is more
In time, more special, sensitiveer, it can be realized the early diagnosis of osteoarthritis, to reduce the death rate of osteoarthritis.
Detailed description of the invention
Fig. 1 shows the expression using QPCR detection PAEP gene in synovial cell；
Fig. 2 shows the expression using Western blot detection PAEP albumen in synovial cell；
Fig. 3 shows the transcription situation using QPCR detection PAEP gene；
Fig. 4 shows the overexpression situation using Western blot detection PAEP albumen；
Fig. 5 shows synovial fluid to the proliferation function of OA fibroblast-like synoviocyte；
Fig. 6 shows PAEP gene overexpression to the inhibiting effect of the OA fibroblast-like synoviocyte proliferation under incentive condition；
Fig. 7 shows the inhibiting effect that PAEP gene overexpression is proliferated OA fibroblast-like synoviocyte.
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.
The differential expression of PAEP gene in 1 OA synovial tissue of embodiment and normal synovial tissue
The synovial tissue of 60 OA patients is from BJ Union Hospital's orthopaedics row knee prosthesis or villusectomy
Patient OA.Case used meets the diagnostic criteria about OA of Altam proposition.40 normal synovial tissues are coordinated from Beijing
Hospital orthopedics are trauma surgery patient synovial tissue of joint.Supernatant, -80 DEG C of storages are taken after OA patient's synovial fluid (SF) high speed centrifugation
It deposits stand-by.Clinical sample used in this research know to patient and informs and pass through through Ethics Committee, the court.
1, the detection of PAEP gene transcription level
1.1 synovial tissue's cell injuring models
After the synovial tissue of sterile acquisition is cleaned with PBS, about 1mm x1mm x is cut into repeatedly with aseptic operation scissors
The tissue block of 1mm filters after 37 DEG C of clostridiopetidase A II (0.5mg/ml), digestion 2h is added through 200 mesh gauzes, after supernatant is removed in centrifugation,
Cell is resuspended in DMEM culture solution, is placed in 37 DEG C, 5%CO2Culture in cell incubator.When cell grows up to shuttle shape and in blocks
Afterwards, secondary culture is carried out.After cell reached for 3 generation, it is separately added into mouse anti human CD3, CD14, CD19 and PE of FITC label
The mouse anti human CD11b of label is marked, flow cytomery identification.Above-mentioned 4 kinds of labels be negative cell be at
Fiber-like synovial cell (Fibroblast-like Synoviocytes, FLS) is used for this research.
1.2 Total RNAs extraction
OA patient synovial tissue cell and normal is extracted using the tissue/cell total RNA extraction reagent box of QIAGEN company
The total serum IgE of synovial tissue's cell.
1.3 reverse transcription
The reverse transcription of RNA is carried out using the Reverse Transcriptase kit of TAKARA company.
1.4QPCR
(1) design of primers
QPCR amplimer is designed according to the coded sequence of PAEP gene and GAPDH gene in Genbank, by the raw work in Shanghai
The synthesis of biotechnology Services Co., Ltd.Specific primer sequence is as follows:
PAEP gene:
Forward primer is 5 '-GCTGTGTTGAGAAGAAGGT-3 ' (SEQ ID NO.3)；
Reverse primer is 5 '-TTCGCCACCGTATAGTTG-3 ' (SEQ ID NO.4),
GAPDH gene:
Forward primer is 5 '-TTTAACTCTGGTAAAGTGGATAT-3 ' (SEQ ID NO.5)；
Reverse primer is 5 '-GGTGGAATCATATTGGAACA-3 ' (SEQ ID NO.6).
(2) PCR reaction system is prepared according to table 1:
Wherein, SYBR Green polymerase chain reaction system is purchased from Invitrogen company.
1 PCR reaction system of table
| Reagent | Volume |
| Forward primer | 1μl |
| Reverse primer | 1μl |
| SYBR Green polymerase chain reaction system | 12.5μl |
| Template | 2μl |
| Deionized water | Supply 25 μ l |
(3) PCR reaction condition: 95 DEG C of 10min, (95 DEG C of 15s, 60 DEG C of 60s) * 46 circulations.Using SYBR Green as
Fluorescent marker carries out PCR reaction on Light Cycler fluorescence quantitative PCR instrument, true by melt curve analysis analysis and electrophoresis
Determine purpose band, Δ Δ CT method carries out relative quantification.
1.5 statistical method
Experiment is completed according to being repeated 3 times, and result data is indicated in a manner of mean+SD,
Using SPSS13.0 statistical software come for statistical analysis, difference between the two is examined using t, it is believed that is had as P < 0.05
It is statistically significant.
1.6 result
As a result as shown in Figure 1, compared with normal synovial tissue, PAEP gene expression amount in Osteoarthritic Synovium histocyte
It significantly reduces, difference has statistical significance (P < 0.05).
2, the detection of PAEP gene expression -- protein level
2.1 steps: the detection of PAEP protein level is carried out using conventional western blot.
2.2 statistical procedures
The gray value of protein band is analyzed using Image J software, using β-actin as internal reference, by PAEP albumen
The gray value of band is normalized.Experiment be all to be completed according to being repeated 3 times, result data be all with average value ±
The mode of standard deviation indicates, using SPSS13.0 statistical software come for statistical analysis, difference between the two is examined using t
It tests, it is believed that there is statistical significance as P < 0.05.
2.3 result
As a result as shown in Fig. 2, compared with normal synovial tissue, the expression of PAEP albumen in Osteoarthritic Synovium histocyte
Amount significantly reduces, and difference has statistical significance (P < 0.05).
The building of 3 PAEP gene expression plasmid of embodiment
1, the overexpression of PAEP gene
The building of 1.1PAEP expression vector
Amplimer is designed according to the coded sequence (as shown in SEQ ID NO.1) of PAEP gene, primer sequence is as follows: just
It is 5 '-ATGCTGTGCCTCCTGCTCA-3 ' (SEQ ID NO.7), reverse primer 5 '-to primer
ACGGCTCTTCCATCTGTTTCA-3'(SEQ ID NO.8).From at Human fetal spleen cDNA library (clontech company, article No.:
638831) coded sequence of the PAEP gene of amplification overall length, above-mentioned cDNA sequence are bis- through restriction enzyme BamHI and XhoI in
It is inserted into the eukaryotic expression vector pcDNA3.1 through restriction enzyme BamHI and XhoI double digestion, connects after digestion
The recombinant vector pcDNA3.1-PAEP of acquisition is used for subsequent experimental.
1.2 transfection
OA fibroblast-like synoviocyte is divided into two groups, respectively control group (transfection pcDNA3.1 empty carrier) and PAEP
Overexpression group (transfection pcDNA3.1-PAEP).The transfection of carrier is carried out using liposome 2000, specific transfection method is according to explanation
The instruction of book carries out.The working concentration of pcDNA3.1 empty carrier and pcDNA3.1-PAEP are 0.5 μ g/ml.
1.3QPCR detection
Specific steps are the same as embodiment 2.
1.4Western detection
Specific steps are the same as embodiment 2.
2, result
As shown in figure 3, being transfected in the cell of pcDNA3.1-PAEP compared with the cell of transfection pcDNA3.1 empty carrier
The mRNA level in-site of PAEP significantly raises, and difference has statistical significance (P < 0.05)；As shown in figure 4, empty with transfection pcDNA3.1
The cell of carrier is compared, and the protein level for transfecting PAEP in the cell of pcDNA3.1-PAEP significantly raises, and difference has statistics
Meaning (P < 0.05).
OA fibroblast-like synoviocyte proliferation experiment under 4 incentive condition of embodiment
1, OA fibroblast-like synoviocyte proliferation experiment under incentive condition
1.1 step
OA fibroblast-like synoviocyte is inoculated in 96 porocyte culture plates, every hole 2x103The μ l of the hole cells//200,37
DEG C, 5%CO2It after incubator is incubated for 12 hours, is added in synovial fluid (1:5 dilution), continues to be incubated for 24 hours, be added3H-TdR(1μ
The hole Ci/), it is further cultured for 24 hours, collects cell, add liquid scintillation solution, β calculating instrument detects cpm value.
1.2 statistical method
Experiment is completed according to being repeated 3 times, and result data is indicated in a manner of mean+SD,
Using SPSS13.0 statistical software come for statistical analysis, difference between the two is examined using t, it is believed that is had as P < 0.05
It is statistically significant.
1.3 result
As a result as shown in figure 5, compared with the control group that synovial fluid stimulation is not added, the FLS proliferation for adding synovial fluid to stimulate is obvious
Accelerate, difference has statistical significance (P < 0.05).
The influence that 5 PAEP gene overexpression of embodiment is proliferated OA fibroblast-like synoviocyte under incentive condition
1, step
Cell culture and plasmid transfection are carried out after transfection 12 hours according to the method for embodiment 3, and (1:5 in synovial fluid is added
Dilution), continue to be incubated for 24 hours, be added3H-TdR (1 hole μ Ci/), is further cultured for 24 hours, collects cell, adds liquid scintillation solution, β
Calculating instrument detects cpm value.
2, statistical method
Experiment is completed according to being repeated 3 times, and result data is indicated in a manner of mean+SD,
Using SPSS13.0 statistical software come for statistical analysis, the difference between PAEP gene overexpression group and control group uses t
It examines, it is believed that there is statistical significance as P < 0.05.
3, result
As a result such as Fig. 6 is shown, compared with the cell of transfection pcDNA3.1 empty carrier, transfection pcDNA3.1-PAEP is inhibited
Cell Proliferation under synovial fluid stimulation, difference have statistical significance (P < 0.05).
The influence that 6 PAEP gene overexpression of embodiment is proliferated OA fibroblast-like synoviocyte
1, step
Cell culture and plasmid transfection is carried out after transfection 12 hours according to the method for embodiment 3 to be added3H-TdR(1μCi/
Hole), it is further cultured for 24 hours, collects cell, add liquid scintillation solution, β calculating instrument detects cpm value.
2, statistical method
Experiment is completed according to being repeated 3 times, and result data is indicated in a manner of mean+SD,
Using SPSS13.0 statistical software come for statistical analysis, the difference between PAEP gene overexpression group and control group uses t
It examines, it is believed that there is statistical significance as P < 0.05.
3, result
As a result such as Fig. 7 is shown, compared with the cell of transfection pcDNA3.1 empty carrier, transfection pcDNA3.1-PAEP cell increases
It grows and obviously slows down, difference has statistical significance (P < 0.05).
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.
Claims (7)
1. the application of people PAEP gene and its expression product in the product of preparation diagnosis osteoarthritis, which is characterized in that described
People PAEP gene and its expression product refer to PAEP gene and its expression product in people synovial tissue cell.
2. application according to claim 1, which is characterized in that the product include: by RT-PCR, real-time quantitative PCR,
Immune detection, in situ hybridization or chip detect the expression of PAEP gene and its expression product to diagnose the production of osteoarthritis
Product.
3. application according to claim 2, which is characterized in that the product with RT-PCR diagnosis osteoarthritis at least wraps
Include the primer of a pair of of specific amplified PAEP gene；The product with real-time quantitative PCR diagnosis osteoarthritis includes at least a pair
The primer of specific amplified PAEP gene；It is described with immune detection diagnosis osteoarthritis product include: and PAEP protein-specific
In conjunction with antibody；The product in situ hybridization diagnosis osteoarthritis includes: the spy with the nucleic acid array hybridizing of PAEP gene
Needle；The product with chip diagnosis osteoarthritis includes: protein chip and genetic chip；Wherein, protein chip include with
The antibody that PAEP protein-specific combines, genetic chip includes the probe with the nucleic acid array hybridizing of PAEP gene.
4. application according to any one of claim 1-3, which is characterized in that the product includes chip, kit.
5. application according to claim 4, which is characterized in that the chip includes genetic chip, protein-chip；It is described
Genetic chip includes solid phase carrier and the oligonucleotide probe for being fixed on solid phase carrier, and the oligonucleotide probe includes being used for
Detect the oligonucleotide probe for PAEP gene of PAEP gene transcription level；The protein-chip include solid phase carrier with
And it is fixed on the specific antibody of the PAEP albumen of solid phase carrier.
6. application according to claim 4, which is characterized in that the kit includes that gene detecting kit and albumen are exempted from
Epidemic disease detection kit；The gene detecting kit includes the reagent for detecting PAEP gene transcription level；The albumen is exempted from
Epidemic disease detection kit includes the specific antibody of PAEP albumen.
7. application according to claim 6, which is characterized in that the reagent include for PAEP gene primer and/or
Probe.
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