CN110277138B - Method for detecting susceptibility gene of hormonal femoral head necrosis - Google Patents
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Abstract
The invention discloses a method for detecting a susceptibility gene of hormonal femoral head necrosis, which comprises the following models: f ═ human genome locus rs2305027 score + locus rs34433978 score + locus rs3833221 score + locus rs143471015 score, where F is the hormonal risk factor for femoral head necrosis; when F is more than or equal to 5, the risk of hormonal femoral head necrosis is high; when F is less than or equal to 4, the risk is low for the hormonal femoral head necrosis, and the risk is positively correlated with the fraction. The detection method can be used for a doctor to evaluate whether a patient is suitable for treatment by using the glucocorticoid medicaments and estimate the risk of femoral head necrosis, so that clinical use of the glucocorticoid medicaments is guided, and accurate medical treatment is implemented.
Description
Technical Field
The invention belongs to the field of gene detection, particularly relates to a method for detecting a susceptibility gene of hormonal femoral head necrosis, and also relates to a kit for detecting the susceptibility gene of the hormonal femoral head necrosis.
Background
Femoral head necrosis (Osteoneprosis of the femoral head, ONFH), also known as Avascular necrosis (AVN), refers to a local blood circulation disorder in the femoral head caused by various causes, resulting in hypoxia, shrinkage and death of bone cells (ZALAVRAS C G, LIEBERMAN J R. Osteoneprosis of the femoral head: evaluation and treatment. J Am Acad ortho Surg,2014,22(7): 455-) 464.), causing hip joint pain and movement disorders. Among the many causes of ONFH, steroid-induced Femoral Head necrosis (SiONFH) is considered to be one of the most important types of ONFH other than traumatic ONFH (MONT M A, CHERIAN J J, SIERRA R J, et al. There are several hypotheses about the pathological mechanism of SiONFH, including blood supply disorder of bone microcirculation, blood coagulation disorder, differentiation disorder of mesenchymal stem cells, lipid metabolism disorder, etc., but individual differences in susceptibility to SiONFH among different patients cannot be completely explained. Ono et al (Ono K, TOHJIMA T, KOMAZAWA T. skin factors of avascular minor of the ferromagnetic head in tissues with high-level-systemic lupus erythematous lipid free of clin ortho ps rei, 1992(277) 89-97) followed 62 patients with Systemic Lupus Erythematosus (SLE) who received high-dose glucocorticoid therapy, but had not found significant correlation between daily dose, cumulative dose and duration of use of the hormone with the occurrence of nfh, and even patients with the occurrence of necrosis with low-dose hormone, the susceptibility of this group to sion nfh could not be explained from hormonal amounts, and genetic factors could be present. Therefore, genetic susceptibility is a further research hotspot for sion nfh. Even with similar hormonal regimens in the same primary disease, there are still individual differences in the incidence of sion fh. Yang et al (LAUSTEN G S, JENSEN J S, OLGAARD K.Neocross of the ferromagnetic head after transfer. acta ortho Scand,1988,59(6):650-654.) found that C3435T or G2677T variation of the MDR1 gene could be involved in the development of SiONFH in SLE patients by studies in SLE patients; however, the consistency is not found to be high by the verifications in the different queues. At present, the genetic susceptibility site of any SiONFH is not defined. Single Nucleotide Polymorphisms (SNPs) are the most common DNA sequence variations. This is defined as a single nucleotide variation in the genome, which may be caused by a transition (transition) or transversion (transition) of a single base, or by an insertion or deletion of a base. The previous research fails to screen genetic differences represented by SNP in a high-throughput broad-spectrum manner, so that the obtained susceptibility sites are very limited, the verification rate in different populations is low, and valuable references cannot be provided for clinical diagnosis. With the development of the genetics research technology, the cost of a new generation sequencing technology (NGS) is gradually reduced, the precision is gradually improved, the depth and the breadth of genetics detection are continuously increased, and a larger amount of genetic susceptibility sites can be found, so that the pathogenesis of SiONFH is revealed, and a more refined SiONFH diagnosis and treatment scheme is provided.
Disclosure of Invention
To date, there is no literature report that high throughput sequencing techniques are used prospectively to detect genes or gene sets associated with hormonal femoral head necrosis and to optimize or select treatment regimens accordingly. In order to fill the blank of the clinical technology, achieve the purposes of accurate medical treatment, ensuring the administration accuracy of glucocorticoid users and improving the treatment success rate, the inventor combines clinical practice and gene sequencing technology to find some hormone femoral head necrosis susceptibility genes, and establishes a hormone femoral head necrosis susceptibility gene detection model based on the polymorphism of the susceptibility genes, wherein the model is a mathematical model and can be used for evaluating whether a certain patient is suitable for treatment by glucocorticoid so as to reduce the risk of femoral head necrosis of the patient. Specifically, the present invention includes the following technical solutions.
A method for detecting a hormone femoral head necrosis (SiONFH) susceptibility gene comprises the following models: f ═ human genome site rs2305027 score, site rs34433978 score, site rs3833221 score and site rs143471015 score, wherein F is the risk coefficient of hormonal femoral head necrosis (sion fh) and is positively correlated with the risk of hormonal femoral head necrosis (sion fh).
In the scoring mode of each locus, the scoring standard of locus rs2305027 is as follows: rs2305027CC, i.e., the two alleles are both C base at the site, and is marked as 0 point; rs2305027CT, namely the C base of one of the two alleles at the site is mutated into a T base, and the score is 1; rs2305027TT, namely the C base of the two alleles at the site is mutated into T base, and the result is marked as2 points;
the scoring criteria for site rs34433978 were: rs34433978GG, i.e., the two alleles are G bases at the locus and are marked as 0 point; rs 34433978G-loss of G base at this site for one of the two alleles, scored as 1 point; rs34433978- -i.e., the G bases of both alleles at this site were lost and scored as2 points;
the scoring criteria for site rs3833221 were: rs 3833221-i.e., no base insertion at this site of the two alleles, and is marked as 0 point; rs 3833221C-one C base is inserted into one of the two alleles at the position, and the mark is 1 point; rs3833221CC, namely a C base is inserted into the site of the two alleles respectively and is marked as2 points;
the scoring criteria for site rs143471015 were: rs143471015GG, namely the G base of the two alleles at the site, and is marked as2 points; rs 143471015G-i.e. the G base deletion of one of the two alleles at this position, is scored as 1 point, rs 143471015-i.e. the G base deletion of both alleles at this position, is scored as 0 point.
In the model, when F is more than or equal to 5, the risk of the hormonal femoral head necrosis is high, and glucocorticoid medicaments are indicated to be used by the patient with caution clinically, and the measures such as dosage reduction, use time and the like are taken; when F is less than or equal to 4, the risk of the hormonal femoral head necrosis is low, which indicates that the patient has low possibility of necrosis by using glucocorticoid medicaments clinically.
Since the model is a mathematical model, it may be input to an information processing module of a gene assaying device or to an information processing module of a blood assaying device.
In one embodiment, the aforementioned hormonal femoral head necrosis susceptibility gene detection model can be inputted into an intelligent medical system or into a computer of a doctor.
Another aspect of the present invention is to provide a kit for detecting a susceptibility gene to hormonal femoral head necrosis, which includes primer sequences (or primer pair sequences) for detecting genes rs2305027, rs34433978, rs3833221 and rs143471015, respectively, so as to be used for amplification and subsequent sequencing detection of the above-mentioned sites.
In a preferred embodiment, the primer pair for detecting the gene rs2305027 is a forward primer SEQ ID NO. 1 and a reverse primer SEQ ID NO. 2; the primer pair for detecting the gene rs34433978 is a forward primer SEQ ID NO. 3 and a reverse primer SEQ ID NO. 4; the primer pair for detecting the gene rs3833221 is a forward primer SEQ ID NO. 5 and a reverse primer SEQ ID NO. 6; the primer pair for detecting the gene rs143471015 is a forward primer SEQ ID NO. 7 and a reverse primer SEQ ID NO. 8.
The amplification primer sequences for detecting the genes rs34433978, rs2305027, rs143471015 and rs3833221 by PCR are not limited to the above sequences SEQ ID NOS: 1-8, and obviously include any other suitable primers.
When the kit is used for carrying out PCR amplification on the gene, a PCR reaction system can be as follows: forward primer, 10 μ M, 1 μ L; reverse primer, 10. mu.M, 1. mu.L; 2xTaq Master Mix, 10. mu.L; DNA sample, 1. mu.g; add ddH2O to 20 μ L, and PCR reaction conditions were: 5min at 95 ℃; (95 ℃ 30sec, Tm ℃ 30sec, 72 ℃ 1 min). times.35 cycles; 7min at 72 ℃; keeping the temperature at 4 ℃.
When the kit is used for carrying out gene detection on target people needing SiONFH risk assessment, various existing methods can be adopted to collect whole genome samples (including saliva, blood, body fluid and the like). And designing amplification primer sequences aiming at the SNP sites rs34433978, rs2305027, rs143471015 and rs3833221, carrying out PCR reaction, and carrying out Sanger method sequencing (or other reliable genetic methods) on the amplification products so as to determine the polymorphic state of each susceptibility site.
After the polymorphism state of the susceptibility site is used as a variable and input into the hormone femoral head necrosis susceptibility gene detection model, the clinical risk coefficient of the patient with SiONFH can be calculated.
The hormone femoral head necrosis susceptibility gene detection model is a comprehensive diagnosis model containing 4 variables, or a risk assessment model, and is a mathematical model for glucocorticoid administration risk prediction, which can be input into an intelligent medical system in the form of computer software (computer program) or a computer of a doctor to help the doctor judge the clinical risk of a femoral head necrosis patient suffering from SiONFH, evaluate whether the femoral head necrosis patient is suitable for treatment by glucocorticoid, guide the clinical use of glucocorticoid drugs, and implement precise medical treatment.
The hormone femoral head necrosis susceptibility gene detection model provided by the invention is verified in clinical practice of Zhongshan Hospital affiliated to the university of Compound Dane, and based on retrospective research results of 38 samples (including 23 cases of necrosis and 15 cases of control samples), the area under the ROC curve calculation curve (AUG) is drawn, and the results show that the AUC of the model is 0.835, the sensitivity (or called sensitivity) is 87%, the specificity (or called specificity) is 80%, and the clinical guidance significance is achieved.
Drawings
Fig. 1 shows ROC curves for validation of the prediction model constructed in the present invention, where AUC is 0.835, sensitivity 87%, specificity 80%.
Detailed Description
The invention looks at the individual difference of susceptibility of different patients to steroid-induced femoral head necrosis (SiONFH) from the genetic perspective, and through screening a large number of genes possibly related to SiONFH in the human genome, at least 4 genomic loci rs2305027 (related to a functional gene COLEC12), rs34433978 (related to a functional gene CYP2B6, CYP2A13), rs3833221 (related to a functional gene CYP2F1) and rs143471015 (related to a functional gene SLFN12L) are highly related, and the mutation of individual bases of the genes form gene polymorphism, and the influence of the polymorphism (including the number of base mutations and the mutation type) on SiONFH is different. Based on this polymorphism, a mathematical model was constructed using these 4 loci as variables.
According to the mathematical model, the clinical risk of glucocorticoid-related femoral head necrosis of patients treated by glucocorticoid can be evaluated, and the clinical use of glucocorticoid medicaments can be guided. That is, the present invention proposes a solution to the problem of clinical practice having difficulty in accurately assessing whether a particular patient is eligible for treatment with a glucocorticoid.
It will be apparent to those skilled in the art that the hormonal femoral head necrosis susceptibility gene testing model can also be in the form of other mathematical models.
Herein, the terms "genomic site", "site" or "SNP site" referring to a site in the human genome associated with sion fh, are intended to have the same meaning and may be used interchangeably, especially referring to a mutable, i.e. mutable site, which may be associated with hormonal femoral head necrosis (sion fh), such as steroid-induced femoral head necrosis.
The mathematical model of the present invention can be input to the information processing module of a gene testing device, such as a gene sequencer, or to the information processing module of a blood testing device, by programming, in the form of a mathematical software package; or can be input into a cloud server to form a shareable intelligent medical system or into a computer of a doctor, and the operation of the equipment can generate a guidance suggestion of the risk of the patient suffering from the hormonal femoral head necrosis.
In the mathematical model of the present invention, in order to indicate that the polymorphism of the gene, that is, the mutation type of the allele, is different, a representation is adopted in which a symbol or a base (selected from A, C, G, T) is appended to the genomic locus rs2305027, rs34433978, rs3833221, rs143471015 as a suffix. The numbering format is a way of internationally universal tagging of SNPs (single nucleotide polymorphisms), and reference SNP numbering is summarized and confirmed by NCBI (national center for Biotechnology information in the United states), by which numbering can be conveniently performed in NCBI database: (http://www.ncbi.nlm.nih.gov/) The related information of the SNP is searched. The symbols or bases (A, C, G, T, -) added after rs numbering represent five meanings, i.e., adenine, cytosine, guanine, thymine and abasic, respectively. The two symbols or the combination of bases represent the base states of the two alleles in the diploid organism cell. If "TT" represents T bases at both alleles at the corresponding positions; "CT" means that at the corresponding positions in both alleles, one is a C base and one is a T base; "G-" represents that one of the two alleles at the position is G base, the other one is vacant, and specifically represents insertion mutation or deletion mutation, and the position is searched and identified in a database according to rs number.
The locus rs2305027 refers to the mutation of 334742 th C base of chromosome 18 into T base. The site is positioned in the exon region of the Colec12 gene, and can cause the function abnormality of the protein coded by the gene. Due to the existence of alleles, the rs2305027 locus has three states of CC, CT and TT, namely CC means that two alleles are C base at the locus, CT means that C base of one of the two alleles at the locus is mutated into T base, and TT means that C base of the two alleles at the locus is mutated into T base. In the scoring mode of the model of the invention, rs2305027CC is scored as 0, rs2305027CT is scored as 1, and rs2305027TT is scored as 2.
rs34433978 refers to deletion mutation of G at 41567532 th position of chromosome 19. The site is located in the intergenic region of CYP2B6 and CYP2A13, which causes frame shift mutation of the segment and seriously affects the gene function. The existence of GG and G < - > at the rs34433978 locus due to the existence of alleles, namely GG means that two alleles have G base at the locus, G < - > means that the G base of one of the two alleles is lost at the locus, and < - > means that the G base of the two alleles is lost at the locus. In the scoring mode of the model, rs34433978GG is marked as 0, rs 34433978G-is marked as 1, and rs 34433978-is marked as 2.
rs3833221 refers to the insertion mutation of the C base at 41622107 th base of chromosome 19. The site is located in the exon region of the CYP2F1 gene, so that the segment is subjected to frame shift mutation, and the function of the protein encoded by the gene is seriously influenced. Due to the existence of alleles, the rs3833221 locus exists, and the three states of C and CC are that-means that two alleles have no base insertion at the locus, C-means that one of the two alleles has a C base insertion at the locus, and CC means that two alleles have a C base insertion at the locus respectively. In the scoring mode of the model, rs3833221- -is marked as 0 point, rs3833221C- -is marked as 1 point, and rs3833221CC is marked as2 points.
rs143471015 refers to 33806842 th base G deletion mutation of chromosome 17, and there are three states of GG and G-, - -at rs3833221, i.e. GG refers to that both alleles are G base at the site, G-refers to that one of the two alleles is G base deletion at the position, and-refers to that both alleles are G base deletion at the position. In the scoring mode of the model, rs143471015- -is marked as 0, rs143471015G- -is marked as 1, and rs143471015GG is marked as 2.
Clinical practices carried out in Zhongshan Hospital affiliated to the university of Compound Dan prove the reliability of the hormone-induced femoral head necrosis susceptibility gene detection model.
In order to make the present invention more comprehensible, embodiments accompanying with the drawings are described in detail below. It will be understood by those skilled in the art that the following examples are only for illustrating the feasibility of the present invention and are not intended to limit the present invention.
All percentages referred to in the examples refer to mass percentages unless otherwise indicated (e.g., explicitly indicated as percentages or ratios).
Example 1 screening of SiONFH-related genes
1.1A prospective observation queue of SiONFH was established according to a prospective nest case control study of Rheumatoid hospitalized patients in Zhongshan Hospital affiliated with university of Redding university from 7/1/2015 to 2019/1/25 (registration number of Chinese clinical trial registration center: ChiCTR-OON-15005843, ethical code B2013-123 (2)). This cohort is included in patients who require the use of glucocorticoids for various rheumatic diseases, at doses up to the conventional high dose (i.e. > 30 mg/day of prednisone equivalent, for 30 days). The cohort was enrolled in 73 eligible patients, all followed by the Han nationality. The follow-up period is 2 years, and 64 patients successfully complete follow-up, wherein 6 patients with necrosis. 6 persons in control case are selected according to the mixed factor pairing of primary disease, sex, age, etc., genome DNA is extracted from blood samples of 12 patients and whole exon sequencing is carried out, 9 different sites are selected, wherein the sites are rs2305027, rs562038978, rs34433978, rs3833221, rs111896385, rs61758879, rs2708381, rs143471015, rs2277969, and 9 functional genes are involved (COLEC12, FMN2, CYP2B6, CYP2A13, CYP2F1, ARHGAP24, TAS2R46, SLFN12L and COL5A 3). The types of variation are related to nonsynonymous mutations, frameshift mutations, nonsense mutations and fragment deletions.
1.2 to further verify the correlation between the above genes and SiONFH, we established a retrospective validation cohort containing 38 samples (23 necrotic samples, 15 controls). The 9 differential sites are detected by Sanger sequencing in a validation queue, and the typing results show that the OR values (odds ratios) of the four sites of rs34433978, rs2305027, rs3833221 and rs143471015 are respectively 19, 15.7, 9.1 and 0.2(p is all less than 0.1), which indicates that the 4 gene sites are highly related to SiONFH.
1.3 indications for inclusion in patients at the time of gene screening: glucocorticoid drugs are required for various reasons, at daily doses greater than 30mg (prednisone equivalent), and are expected to be used for more than 1 month in patients between 18 and 65 years of age. It should be noted that if the patient is associated with other well-defined high risk factors for femoral head necrosis, such as alcoholism and hip primary disease like femoral neck fracture, the patient should be treated at high risk regardless of the evaluation result.
Biological sample retention and genome DNA extraction: genomic samples are collected using either existing saliva collection methods (e.g., saliva collection tubes) or whole blood collection methods (e.g., conventional venous blood collection). And (4) storing the collected sample at-80 ℃ until unified extraction and detection. As for the extraction of genomic DNA, QIAamp tissue DNA extraction kit (QIAGEN, USA) or the like can be used.
1.4 PCR primers shown in Table 1 below were designed for 4 gene loci.
TABLE 1 amplification primers for each Gene site
1.5 PCR amplification
And (3) PCR reaction system: forward primer, 10 μ M, 1 μ L; reverse primer, 10. mu.M, 1. mu.L; 2xTaq Master Mix, 10. mu.L; DNA sample, 1. mu.g; add ddH2O to 20. mu.L.
And (3) PCR reaction conditions: 5min at 95 ℃; (95 ℃ 30sec, Tm ℃ 30sec, 72 ℃ 1 min). times.35 cycles; 7min at 72 ℃; keeping the temperature at 4 ℃.
1.6 DNA sequencing by dideoxy chain nucleic acid termination method (Sanger method):
1) sample preparation: mixing the DNA sample with fluorescein, and adding the mixture into an upper sample plate;
2) running an ABI 3730 sequencer to perform electrophoresis on the DNAs with different sizes in the sample, so that the DNAs are separated according to the molecular size and pass through a detection window;
3) upon passage through the window, a fluorescein signal is detected, and sequencing data is collected and presented.
Example 2 creation of a mathematical model
2.1 based on the results of example 1, the above 4 loci were organized into score tables, and the statistical data shown in Table 2 below were summarized.
TABLE 2 clinical risk score of SiONFH at each gene locus
In Table 2, rs34433978 is taken as an example, "GG" represents that the corresponding positions of both alleles are G bases, "G-" represents that one of the alleles has a base deletion, "and" - "represents that both alleles have a base deletion; taking rs2305027 as an example, the "CC" represents that the corresponding positions of two alleles are both C bases, the "CT" represents that the alleles are respectively C bases and T bases, and the "TT" represents that the corresponding positions of two alleles are both T bases; and so on.
2.2 combining with clinical practice, according to Table 2 above, the scores of each site are added to total score, with a high risk score of not less than 5 and a low risk score of not more than 4, and the risk score increases with increasing score. If a patient scored as high risk is recommended to follow up on hip magnetic resonance at regular 1, 3, 6 months after hormone administration, the cumulative amount of hormone is minimized, or, if possible, the amount of hormone is reduced below the inclusion condition of the present invention (30mg/d prednisone equivalent).
Based on the method, a hormone femoral head necrosis susceptibility gene detection model is established, which comprises the following steps: f ═ locus rs2305027 score, locus rs34433978 score, locus rs3833221 score and locus rs143471015 score, wherein F is the risk coefficient of steroid-induced femoral head necrosis (sion nh), and when F is more than or equal to 5, the risk is high risk of hormonal femoral head necrosis; when F is less than or equal to 4, the risk of hormonal femoral head necrosis is low.
Obviously, the detection model can be further expanded to include scoring of other mutation classes, as will be appreciated by those skilled in the art.
At present, no literature reports that similar models can accurately predict the occurrence of SiONFH exist, and the combined diagnosis model or risk assessment model of a plurality of genes provided by the invention provides important reference for predicting the occurrence of SiONFH. The all cause necrosis rate in the look-ahead queue was 8.2%. The risk assessment model was 23.8% for high risk patients and 0.5% for low risk patients (p < 0.05). The results show that the model has higher accuracy and clinical guidance significance.
Combining the femoral head necrosis classification diagnosis result of the example 1 and the mathematical model, drawing an ROC curve, calculating the area under the curve (AUG), and displaying that the AUC of the four-site combined diagnosis model is up to 0.835, the sensitivity is 0.87 and the specificity is 0.80. As shown in fig. 1.
Example 3 clinical case of application of the mathematical model
Application example the prediction model established in example 2 was verified.
Case one: SiONFH patients
A 38 year old female patient diagnosed with adult still's disease. The patient was typed for 60mg/d glucocorticoid: rs2305027CT, rs34433978- -, rs3833221CC, rs143471015GG, SiONFH risk score of 7, classified as high risk. MRI after 1 month showed bilateral femoral head necrosis.
Case two: non-SiONFH
A53 year old female patient for diagnosing systemic lupus erythematosus. The patient was typed for glucocorticoid at 40 mg/d: rs2305027CC, rs34433978GG, rs3833221- -, rs143471015-G, SiONFH risk score of 1, classified as low risk. Femoral head necrosis is not seen in 3 years of follow-up.
The technical scheme of the invention is verified by the embodiment, the reliability of the prediction model of the invention is proved, and accurate prejudgment can be provided for distinguishing patients who use glucocorticoid medicaments from patients who use the glucocorticoid medicaments cautiously. It should be noted that various changes or modifications made by those skilled in the art without departing from the spirit of the invention shall also fall within the scope of the invention.
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Claims (8)
1. A kit for detecting a hormone-induced femoral head necrosis susceptibility gene, which comprises primer sequences for detecting human genome sites rs2305027, rs34433978, rs3833221 and rs143471015 respectively, so as to be used for amplification and subsequent sequencing detection of the sites, and comprises the following models: f ═ human genome locus rs2305027 score, locus rs34433978 score, locus rs3833221 score and locus rs143471015 score, wherein F is a hormonal femoral head necrosis risk coefficient and is in positive correlation with the hormonal femoral head necrosis risk, and wherein F is a hormone-induced femoral head necrosis risk coefficient and is in positive correlation with the hormonal femoral head necrosis risk
The scoring criteria for locus rs2305027 are: rs2305027CC, i.e., the two alleles are both C base at the site, and is marked as 0 point; rs2305027CT, namely the C base of one of the two alleles at the site is mutated into a T base, and the score is 1; rs2305027TT, namely the C base of the two alleles at the site is mutated into T base, and the result is marked as2 points;
the scoring criteria for site rs34433978 were: rs34433978GG, i.e., the two alleles are G bases at the locus and are marked as 0 point; rs 34433978G-loss of G base at this site for one of the two alleles, scored as 1 point; rs34433978- -i.e., the G bases of both alleles at this site were lost and scored as2 points;
the scoring criteria for site rs3833221 were: rs 3833221-i.e., no base insertion at this site of the two alleles, and is marked as 0 point; rs 3833221C-one of the two alleles has a C base inserted into the site, and the score is 1; rs3833221CC, namely, a C base is inserted into each of the two alleles at the site, and the score is 2;
the scoring criteria for site rs143471015 were: rs143471015GG, namely the G base of the two alleles at the site, and is marked as2 points; rs 143471015G-one of the two alleles has G base deletion at the position, which is marked as 1 point, and rs 143471015-one of the two alleles has G base deletion at the position, which is marked as 0 point.
2. The kit of claim 1, wherein when F ≧ 5, there is a high risk of hormonal femoral head necrosis; when F is less than or equal to 4, the risk of hormonal femoral head necrosis is low.
3. The kit of claim 1, wherein the model is inputted to an information processing module of a gene assaying device or to an information processing module of a blood assaying device.
4. The kit of claim 1, wherein the model is entered into an intelligent medical system or into a physician's computer.
5. The kit of claim 1, wherein the primer pair for detecting gene rs2305027 is a forward primer of SEQ ID NO. 1 and a reverse primer of SEQ ID NO. 2.
6. The kit of claim 1, wherein the primer pair for detecting the gene rs34433978 is a forward primer of SEQ ID NO. 3 and a reverse primer of SEQ ID NO. 4.
7. The kit of claim 1, wherein the primer pair for detecting the gene rs3833221 is a forward primer SEQ ID NO. 5 and a reverse primer SEQ ID NO. 6.
8. The kit of claim 1, wherein the primer pair for detecting gene rs143471015 is a forward primer SEQ ID NO 7 and a reverse primer SEQ ID NO 8.
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| CN113801931B (en) * | 2020-09-01 | 2024-03-08 | 吉林大学 | CEBPA, PPARgamma, CREBBP gene SNP detection substance related to ONFH risk and application thereof |
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