KR20180040461A - Method of predicting the risk of a rejection in kidney transplant patients using SNPs in BNIP2 gene - Google Patents

Abstract

The present invention relates to a method for predicting an adverse response after organ transplantation using nucleotide sequence polymorphism in a BNIP2 gene site and, more specifically, to a composition for predicting an adverse response after organ transplantation, comprising a probe or primer capable of detecting single nucleotide polymorphism, NCBI refSNP ID: rs754641, of a BNIP2 gene as an active ingredient, a method for detecting single nucleotide polymorphism, NCBI refSNP ID: rs754641, of a BNIP2 gene through sequence analysis from a sample of a patient to provide information required for predicting an adverse reaction after organ transplantation, and a kit for predicting an adverse reaction after organ transplantation. According to the present invention, since the occurrence of an adverse reaction after organ transplantation is changed according to the genetic difference of individuals, such a prediction may be useful for the establishment of differentiated immunosuppression treatment strategies for preventing an adverse reaction and the early detection of an adverse reaction through regular monitoring of a patient group at high risk for an adverse reaction. The kit for the prediction and early detection of an adverse reaction after organ transplantation can contribute to the activation of the organ transplantation industry by improving the clinical and economic outcome of patients.

Description

Methods for predicting rejection after organ transplantation using nucleotide polymorphisms of the BNIP2 gene locus [

The present invention relates to a method for predicting a rejection reaction after organ transplantation using a nucleotide sequence polymorphism of the BNIP2 gene region, and more particularly, to a method for predicting a rejection reaction after transplantation using a probe or a primer capable of detecting NCBI refSNP ID: rs754641 single nucleotide polymorphism of BNIP2 gene as an active ingredient A method for detecting a single nucleotide polymorphism of NCBI refSNP ID: rs754641 BNIP2 gene by analyzing a base sequence from a patient sample to provide information for predicting a rejection reaction after organ transplant including a transplant, And NCBI refSNP ID of BNIP2 gene: rs754641 and rs2306359, or a primer for genotyping. The present invention relates to a kit for predicting a rejection reaction after organ transplantation, comprising a primer for screening a polymorphism selected from the group consisting of NCBI refSNP ID: rs754641 and rs2306359.

Transplantation is defined as a treatment that restores the function of an organs of a terminally ill patient who may lose their life due to the organs of the patient's organs that are no longer functioning and which are difficult to recover by conventional therapies. Approximately 32-75% of patients develop rejection after organ transplantation, of which about 50% occur within 3 months after transplantation, some of whom have lost transplanted grafts due to rejection (Kasiske BL, et al: The evaluation of renal transplantation candidates: Clinical practice guidelines. American Journal of Transplantation: The American Society of Transplantation and the American Society of Transplant Surgeons 2001; 1 Suppl 2: 3-95 ). It is known that early rejection reactions after organ transplantation are mainly caused by antibodies against human leukocyte antigen (HLA), and activation of allogeneic T cells and antigen presenting cells HLA congruity between the transplant donor and recipient (tissue fitness) and the degree of immune activity of the recipient are important factors. However, even if HLA is completely matched, it is estimated that about 20% of the patients have a rejection reaction within 10 years, so that other factors besides HLA are involved (Terasaki, PI: Duration of immunologic and non-immunologic failure in cadaver donor transplants. Clinical transplants 2003; 449-52).

A number of genetic studies have been carried out to investigate the factors and mechanisms involved in rejection reactions as attempts have been made to increase the transplant outcome through prophylactic treatment. In the case of interleukin-10 (IL-10) genotypes, some studies have reported that they are associated with the risk of rejection after renal transplantation, but others have not. In the meta-analysis, there was no statistically significant association between IL-10 1082 GG genotype and rejection. In addition, genetic mutations of IL-4, MCP-1 and TGF-β, which are mainly related to the inflammatory response, have been found to have a significant correlation with rejection occurrence. These studies have attempted to identify various clinical and genetic factors for predicting rejection after kidney transplantation. However, none of these factors have predicted the risk of rejection, The development of new biomarkers that are required to be

In recent years, studies have shown that the development of post-transplant rejection is associated with microRNA, and thus the clinical significance of the 3'-untranslated region (3'-UTR), a target gene-binding site of a specific microRNA, has been attracting attention. With the development of whole exome sequencing technology, genetic polymorphisms in the 3'-UTR region are expected to uncover new genotypes associated with the incidence of rejection.

Numerous papers and patent documents are referenced and cited throughout this specification. The disclosures of the cited papers and patent documents are incorporated herein by reference in their entirety to better understand the state of the art to which the present invention pertains and the content of the present invention.

Therefore, the inventors of the present invention conducted a study to develop a technique for predicting a rejection reaction after organ transplantation, and as a result, by searching the various genetic information of an individual through a full-length exome sequence analysis in a transplant patient group, The present inventors have found that haplotype is closely related to the occurrence of rejection after organ transplantation independently of HLA organization fitness, donor type, age, and sex.

Accordingly, an object of the present invention is to provide a composition for predicting a rejection reaction after organ transplantation, which comprises, as an active ingredient, a probe or primer capable of detecting NCBI refSNP ID: rs754641 single nucleotide polymorphism of BNIP2 gene.

Another object of the present invention is to provide a method for detecting a single nucleotide polymorphism of NCBI refSNP ID: rs754641 BNIP2 gene by analyzing the nucleotide sequence from a patient sample to provide information necessary for predicting rejection reaction after organ transplantation.

Another object of the present invention is to determine the haplotype of the single nucleotide polymorphism site and to identify the haplotype of the BNIP2 gene for predicting the rejection response after organ transplantation in which a specific haplotype is identified as having a high or low risk of rejection after organ transplantation And to provide a method for analyzing single nucleotide polymorphisms.

Another object of the present invention is to provide a kit for predicting rejection response after organ transplantation comprising a screening primer or a genotyping primer selected from the group consisting of NCBI refSNP ID: rs754641 and rs2306359 of BNIP2 gene.

In order to achieve the above object, the present invention provides a composition for predicting a rejection reaction after organ transplantation comprising a probe or a primer capable of detecting NCBI refSNP ID: rs754641 single nucleotide polymorphism of BNIP2 gene as an active ingredient.

In order to achieve the above object, the present invention provides a method for detecting a single nucleotide polymorphism of NCBI refSNP ID: rs754641 BNIP2 gene by analyzing a nucleotide sequence from a patient sample to provide information necessary for predicting rejection reaction after organ transplantation ≪ / RTI >

In order to accomplish still another object of the present invention, there is provided a method for predicting a rejection reaction after organ transplantation in which a haplotype related to the single nucleotide polymorphism site is determined and a specific haplotype is determined to have a high or low risk of rejection after organ transplantation A method for analyzing single nucleotide polymorphism of BNIP2 gene is provided.

In order to accomplish still another object of the present invention, the present invention provides a method for predicting a rejection reaction after organs including a screening primer or a genotyping primer selected from the group consisting of NCBI refSNP ID: rs754641 and rs2306359 of BNIP2 gene For example.

Hereinafter, the present invention will be described in detail.

The term " single nucleotide polymorphism " in the present invention means a genetic change or mutation showing a difference in one nucleotide sequence (A, T, G, C) in the nucleotide sequence of DNA is more than 2% It refers to genetic trait information on which sequences can occur.

The term " next generation sequencing (NGS) " in the present invention refers to a method of dividing a genome into numerous pieces, analyzing the genetic information of each fragment, combining and analyzing the entire nucleotide sequence.

The term 'whole exome sequencing (WES)' in the present invention means sequencing only gene regions involved in protein production as one of the next-generation sequencing methods.

In the present invention, the term 'haplotype' refers to a series of SNPs located on one chromosome at the time of germ cell division, which can be divided into several bundles according to their inheritance relation. In this case, , And this haplotype is a combination of several SNPs present on the same chromosome and exhibits a constant SNP pattern.

The composition of the present invention is characterized by comprising a probe or primer capable of detecting NCBI refSNP ID: rs754641 single nucleotide polymorphism of BNIP2 gene as an effective component for predicting rejection reaction after organ transplantation.

The polymorphic site of the present invention is located in the exon of the BNIP2 gene and is a site specified by rs754641 and rs2306359 as NCBI refSNP ID.

rs754641 is a SNP located in the exon 50 region of BNIP2, located at position 59689391 of chromosome 15, and having G mutated to A. rs754641 can be composed of 8 to 100 consecutive DNA sequences substituted with A at position 258 of the nucleotide sequence of SEQ ID NO: 1.

rs2306359 is SNP located in the exon 149 region of BNIP2 and located at 59689292th position of chromosome 15, and T is a SNP mutated to G. rs2306359 can be composed of 8-100 consecutive DNA sequences substituted with G at the 159th base T of the nucleotide sequence shown in SEQ ID NO: 2.

The analysis of association between the BNIP2 gene polymorphism and rejection after renal transplantation showed that the risk of rejection was decreased with increasing basal polymorphism of rs754641. The more the polymorphism of rs2306359, Of the total population. These results indicate that rs754641 SNP and rs2306359 SNP can be used as a predictor of rejection after organ transplantation.

The term " diagnosis " as used herein includes any type of analysis used to predict or predict the disease or condition of a patient and to determine the risk of developing a pathological particular condition.

And the term " polymorphism " in the present invention means the occurrence of two or more alternative sequences or alternate alleles within a genetically determined population. Preferred polymorphic markers have two alleles that exhibit an incidence of 1% or more, more preferably 10% or 20% or more in selected populations.

The term " single nucleotide polymorphism " in the present invention means a mutation of a DNA nucleotide sequence in which a single nucleotide A, T, C or G in the genome is different in the same species or between chromosome pairs of each individual.

The present invention relates to a base mutant at each SNP position in a DNA base sequence, but also includes a polynucleotide sequence complementary to the aforementioned nucleotide sequence when such SNP base mutation is found in a double-stranded gDNA (genomic DNA) Is interpreted. Thus, the base at the SNP position in the complementary polynucleotide sequence is a complementary base.

The present invention relates to a method for detecting a single nucleotide polymorphism of the BNIP2 gene consisting of NCBI refSNP IDs: rs754641 and rs2306359 through sequence analysis from a patient sample to provide information necessary for prediction of rejection after organ transplantation .

In the present invention, the analysis of the nucleotide sequence of the SNP, i.e., genotyping, can be performed according to various methods known in the art. Sequence analysis can be performed by sequencing, microarray hybridization, allele specific PCR, dynamic allele-specific hybridization (DASH), PCR extension analysis, SSCP But is not limited to, Single Strand Conformation Polymorphism (DGGE), Restriction Fragment Length Polymorphism (PCR-RFLP), Two Dimensional Gene Scanning (TDGS), TOGATM, and the like.

For example, a method using an Affymetrix SNP chip, a Single Strand Conformation Polymorphism (PCR) method, a PCR-SSO (Specification Sequence Oligonucleotide) method, a dot hybridization method, a PCR- (Allele Specific Oligonucleotide) Hybridization Method, TaqMan Method, Pyrosequencing Method, Invader Method, SNaPShot Method (Allele Specific Gene Amplification Method), MassArray Method, Microplate Array Diagonal Gel Electrophoresis Microplate Array Diagonal Gel Electrophoresis (MADG) method, GoldenGate analysis method, SNPlex method and BeadArray method.

In the method of the present invention, a primer for amplifying a nucleic acid molecule or a probe for detecting a nucleic acid molecule may be used for single nucleotide sequence analysis.

The term " primer " as used herein refers to a single strand of oligonucleotides which is hybridized under suitable conditions (presence of four different nucleoside triphosphates and polymerase such as DNA or RNA polymerase) Quot; means acting as a starting point at which to initiate directed DNA synthesis. The suitable length of the primer is determined by the characteristics of the primer to be used, but is usually 15 to 30 bp in length. The primer need not be exactly complementary to the sequence of the template, but should be complementary enough to form a hybrid-complex with the template.

The term " probe " in the present invention means a linear oligomer having a natural or modified monomer or linkage comprising a deoxyribonucleotide and a ribonucleotide that can hybridize to a specific nucleotide sequence. Preferably, the probe is single stranded for maximum efficiency in hybridization. The probe is preferably a deoxyribonucleotide. As the probe used in the present invention, a sequence completely complementary to the sequence including the SNP may be used, but a sequence substantially complementary may be used as long as it does not interfere with the specific hybridization. Suitable conditions for hybridization are described in Joseph Sambrook, et al .: Molecular Cloning, A Laboratory Manual, Cold Spring Harbor Laboratory Press, Cold Spring Harbor, N.Y. (2001) and Haymes BD, et al .: Nucleic Acid Hybridization, A Practical Approach, IRL Press, Washington, D.C. (1985). ≪ / RTI >

The stringent condition used for hybridization can be determined by controlling the temperature, the ionic strength (buffer concentration) and the presence of a compound such as an organic solvent, and the like. This stringent condition can be determined differently depending on the sequence to be hybridized.

Meanwhile, the present invention provides a method for analyzing a single nucleotide polymorphism of the BNIP2 gene family for predicting rejection response after organ transplantation, the method comprising the following steps.

(a) isolating genomic DNA from an individual;

(b) determining the nucleotide sequence of the NCBI refSNP ID: rs754641 and rs2306359 sites;

(c) detecting a nucleotide sequence polymorphism in which G is replaced by A in the case of rs754641 and G is replaced by G in the case of rs2306359;

(d) determining a haplotype with the detected base sequence polymorphism; And

(e) Haplotypes H1 and H2 are high risk for rejection after organ transplantation, and H3 is low risk for rejection after organ transplantation.

In the single base polymorphism analysis method of the present invention, step (a) is a step of isolating nucleic acid from an individual.

The term "nucleic acid" in the present invention encompasses both DNA (gDNA and cDNA) and RNA molecules. Nucleotides which are basic constituent units in nucleic acid molecules include not only natural nucleotides but also analogues analogue).

In the present invention, the nucleic acid molecule can be obtained from various types of " biological sample "isolated from an individual or individual and can be obtained, for example, from tissues, whole blood, serum, plasma, body fluids, urine, Preferably, but not limited to, tissue or whole blood.

The step of obtaining the nucleic acid from the specimen can be carried out by a conventional DNA separation method or RNA isolation and reverse transcription. In the present invention, when the nucleic acid molecule is a genomic DNA (gDNA), the separation of gDNA can be performed according to a conventional method known in the art (see Rogers & Bendich (1994)), GDNA can be isolated using a commercial kit known in the art (e.g., QIAamp DNA Blood Maxi Kit). When the starting material is mRNA, the total RNA is isolated by a conventional method known in the art (see Sambrook, J. et al., Molecular Cloning. A Laboratory Manual, 3rd ed. Cold Spring Harbor Press (2001); Ausubel, F.M. et al., Current Protocols in Molecular Biology, John Willey & Sons (1987); And Chomczynski, P. et al., Anal. Biochem. 162: 156 (1987). The isolated total RNA is synthesized by cDNA using reverse transcriptase. Because the total RNA is isolated from animal cells, it has a poly-A tail at the end of mRNA. CDNA can be easily synthesized using oligo dT primers and reverse transcriptase using such a sequence characteristic (see PNAS USA, 85: 8998 (1988); Libert F, et al., Science, 244: 569 (1989); and Sambrook, J. et al., Molecular Cloning, A Laboratory Manual, 3rd ed. Cold Spring Harbor Press )).

(b) is a step of determining the nucleotide sequence of the NCBI refSNP ID: rs754641 and rs2306359 regions, and the determination of each SNP and nucleotide sequence is as described above.

(c) is a step of detecting a nucleotide sequence polymorphism in which G is replaced with A in the case of rs754641 and G is replaced with G in case of rs2306359 in the determined nucleotide sequence.

(d) is a step of determining a haplotype with the detected base sequence polymorphism. The determination of monoclast is determined according to Table 5 below for rs754641 and rs2306359.

[Table 5]

Step (e) is a step to determine that the haplotype is at high risk for rejection after organ transplantation for H1 and H2, and low risk for rejection after organ transplantation for H3.

The risk of rejection after organ transplantation depends on the function or activity of BNIP2, depending on the genotype determined by the haplotype. .

In the present invention, the term 'organ transplantation' may be a kidney transplantation, a liver transplantation, a heart transplantation, a lung transplantation, a hematopoietic stem cell transplantation, and preferably a kidney transplantation.

In the present invention, 'prediction of rejection after organ transplantation' is related to graft survival rate and patient morbidity and mortality, so that the individual in the present invention may be a recipient who has received a human or organ transplantation. Preferably a recipient who has undergone renal transplantation.

Meanwhile, the present invention provides a kit for predicting rejection reaction comprising a primer for screening of NCBI refSNP ID: rs754641 single nucleotide polymorphism of BNIP2 gene or a primer for genotyping. In addition, for the screening of additional nucleotide sequence polymorphisms in the kits of the present invention, primers for screening of NCBI refSNP ID: rs2306359 nucleotide polymorphism or primers for genotyping may additionally be included.

The kit of the present invention may contain not only primers capable of amplifying to detect the SNP site of the present invention but also reagents necessary for polymerization such as dNTPs, various polymerase and coloring agents.

In one embodiment of the present invention, DNA was obtained from organ transplant patients, and rejection-related SNPs were selected within 1 year after transplantation. The propensity scores of the rejection group and the non - occurrence group were matched. The inclination score matching included the HLA organization fitness, the type of transplant organ donor, and the age and gender of transplant recipient. As a result, mutations were confirmed from a total of 27084 SNPs, and 24 pharmacokinetic genetic mutations including exon, UTR and intron SNP were identified. In addition, the SNPs of the BNIP2 gene, NCBI refSNP ID: rs754641 and rs2306359 were found to be significantly higher than those of the BNIP2 gene. Respectively.

For reference, the above-mentioned nucleotide and protein work can be referred to the following references (Maniatis et al .: Molecular Cloning: A Laboratory Manual, Cold Spring Harbor Laboratory, Cold Spring Harbor, NY (1982); Sambrook, et Inc., San Diego, Calif. (1990)), which is commercially available under the trade designation "Molecular Cloning: A Laboratory Manual, 2d Ed., Cold Spring Harbor Laboratory Press .

Therefore, according to the present invention, the risk of rejection after organ transplantation can be predicted according to the genetic difference of an individual. Therefore, it is possible to personalize the immunosuppressive therapy strategy for prevention of the rejection reaction before the organ transplantation and to be useful for the early diagnosis and treatment of the rejection reaction through careful monitoring after transplantation. As a result, the clinical and economic performance of organ transplant patients can be improved.

FIG. 1 shows the risk of rejection reaction according to the variation of rs754641 of the present invention (additive: GG vs GA vs. AA).
FIG. 2 shows the risk of rejection reaction according to the variation of rs754641 of the present invention (dominant: GG vs GA + AA).
FIG. 3 shows the risk of rejection reaction according to the variation of rs2306359 of the present invention (dominant: TT vs TG + GG).
Fig. 4 shows a haplotype related to prediction of rejection response after organ transplantation invented through NGS (SNP1: rs2306459, SNP2: rs754641).

Hereinafter, the present invention will be described in detail.

However, the following examples are illustrative of the present invention, and the present invention is not limited to the following examples.

< Example  1>

Collection of test subjects and specimens

The subjects of this study were recruited from Seoul National University Hospital, Sungkyunkwan University Hospital, Seoul, Korea. The protocol of this study was approved by the Bioethics Review Committee of the Seoul National University Hospital (IRB No. C-1504-009-662) and all patient information was collected after written consent.

From 2007 to 2013, patients with renal transplantation who were diagnosed with rejection were divided into two groups according to their demographic characteristics (sex, age at transplantation), donor type, HLA histology, The patients who were matched one-to-one by calculating tendency scores were selected and the final control group was selected.

Demographic characteristics of the recruited patients (test group and control group) are shown in Table 1.

Demographic characteristics of patients with and without rejection summary Test group
(n = 40) Control group
(n = 40) P -value Sex, Male ^† 28 (70%) 21 (52.5%) 0.111 Age (yr) ^‡ 43.85 + 16.61 44.20 ± 14.07 0.918 Donor Type ^†
Living donor
Deceased donor
24 (60%)
16 (40%)
24 (60%)
16 (40%) 1,000 HLA organization fitness ^‡ 2.53 + - 1.63 2.28 ± 1.75 0.511 Induction therapy type
IL2-RA
Anti-thymocyte globulin
38 (95%)
2 (5%)
39 (97.5%)
1 (2.5%) 0.562 ^† n (%), ^‡ mean ± SD
IL2-RA, interleukin-2 receptor antagonist

Blood samples for genetic analysis were collected from peripheral blood of 80 kidney transplant patients and DNA was extracted using QuickGene DNA whole blood kit (KURABO, Japan). Nanodrop ^TM (Thermo Fisher Scientific, US) at 260 nm, 280 nm, and 230 nm, and DNA concentration and purity were measured and stored at 80 ° C.

&Lt; Example 2 >

Selection of SNPs associated with rejection after organ transplantation

<2-1> Full-length exome sequence analysis

DNA was isolated, purified, isolated and amplified using a SureSelect Human All Exon + UTR v5 kit (Agilent, USA) to identify the whole exome and untranslated region sequences. Respectively. The sequences were sequenced in Ion Proton using the library, and the sequences were analyzed by annotation of the analyzed sequences using the analysis pipeline of the torrent server. As a result of the quality check (QC), it was confirmed that the mapping ratio of all samples was more than 70% and the coverage depth was more than 90% in 20x or more as a result of Ion reporter software.

<2-2> Collection of diagnostic information for rejection

The evaluation period of rejection reaction was 1 year after renal transplantation. Renal biopsy was performed to diagnose rejection in organ transplant recipients. Rejection reaction occurred when histologically T - cell - mediated acute rejection was diagnosed. Rejection response diagnostic information was collected retrospectively using electronic medical records.

<2-3> Statistical analysis of analyzed genes

In order to determine the difference in SNP-specific rejection incidence among the two groups matched by the propensity score, ANOVA test was performed using linear models for each SNP by coding each phenotype according to heteromutant and homo-mutant status. In the ANOVA analysis, genotypes with a significance level (P-value) of less than 0.001 were selected.

Mutations were confirmed from a total of 27084 SNPs. A total of 24 genetic mutations, including exons, UTRs and intron SNPs, which were significantly associated with rejection, were identified. Based on the review of the literature, two SNPs of exon were ultimately derived as genotypes that could affect clinically. Table 2 shows the information on the rejection-related SNPs, and the frequencies of the rejection patients by genotype of each SNP are shown in Tables 3 and 4.

Rejection-Associated BNIP2 Gene SNP Information rs  number Gene Location start num variant MAF rs754641 BNIP2 exon 50 15: 59689391 G> A 0.48194 rs2306359 BNIP2 exon 149 15: 59689292 T> G 0.244

Number of patients by genotype of BNIP2 rs754641 GG GA AA Rejection incidence group 22 10 8 Rejection incidence group 6 15 19

Number of patients by genotype of BNIP2 rs2306359 TT TG GG Rejection incidence group 18 13 9 Rejection incidence group 32 8 0

< Example  3>

Analysis of single nucleotide polymorphism and determination of haplotype

In order to evaluate the linkage disequilibrium (LD) relationship between the two SNPs selected to be significant through full-length exon sequence analysis, a solid spine of LD analysis was performed using Haploview (Broad Institute, USA) Respectively.

As a result, as shown in FIG. 4, it was confirmed that two SNPs confirmed by the present invention had a haplotype block, and the frequencies are shown in Table 5 below.

Frequency of SNP haplotype blocks Haplotype rs754641 rs2306359 Frequency H1 G T 0.27544 H2 G G 0.24463 H3 A T 0.48194

As described above, according to the present invention, since the rejection reaction after organ transplantation changes depending on the genetic difference of an individual, prediction of the rejection reaction can be made to establish a differentiated immunosuppressive therapy strategy for prevention of rejection and regular monitoring And can be used for early detection of a rejection reaction. These preventive and early diagnosis kits for the development of rejection reactions after organ transplantation can contribute to the activation of the organ transplantation industry by improving the clinical and economic performance of patients.

<110> Seoul National University R & DB Foundation <120> Method of predicting the risk of a rejection in kidney transplant          patients using SNPs in BNIP2 gene <130> NP16-0125 <160> 2 <170> Kopatentin 2.0 <210> 1 <211> 1093 <212> DNA <213> Artificial Sequence <220> <223> BIP2 SNP rs754641 <220> <221> misc_feature <222> (258) <223> R represents G or <400> 1 ccccggagga agccttggcc ccctcgtcct cttcgcccct ccaggccggc gacgtggggc 60 tgacggccag gtcgcaaaaa gcagggccga gcggagcccg ctcccctcgg tcggcggtgg 120 agaccccggc ccaatccccc ggccgcagcg gtacggcgkc ggcggcagca gctgacccgg 180 acrcagtgag aagccccggc ggaagtgata acatcccgac ctcctccggg cgcggcagcc 240 ggcggtggag gcggcggrgg cgtggcggcg gcggaggcgt ggcgtcttct tgaaacccac 300 acagcatccc cccaaccccc tcagacgtca cgggctagcg ctcgcgcctg cactcgcggg 360 gtcggccttc cttgcctgtg ggcggayccg tcgcgtcgac tccttattgg tggcgttact 420 acgcctcaag gcgtttcttc cctctttgct gtccgcaggg acgctgcctg gtagtttcag 480 ttacttttgt tttctttctg cctgccttta ttgatcatta cagtaggtga aattatcgat 540 atcaatata atattgtaat acaataggtg caattatcaa tatcaatatt attacaacag 600 gtgcaattat cagtatcatt acaataggtg caattatcag cgatcaataa tngtgaatta 660 taaaatggat gaaataaggg gaggaggtcg ggcgcggtgg tttacgcttg taatcccagc 720 actttgggag ggcgaggcgg gtggattacc ggaggtcaag agctcgagac cagcctggcc 780 aacatggtga cacctcgtct ctactaaaca aacaaaaatt agctgggcgt ggtggcgcgc 840 gtctgtaatc ccaactactc aggaggcgag ggcggaggcg gaggcggagt tggagtttgc 900 agtgagccga gatcgtgcca ttgcactcca gcttgggcaa cagagcaaaa actccgtctc 960 aaaaaaagg aaaaaaaaaaaaaaaaaaaaaaaaaaaaaaaaaaagaaataaggcga 1020 ggaaaaatat ccgaggcatc attttgggga gttaggcctc agaagcgacc ttagagtttg 1080 gtaatattkg agt 1093 <210> 2 <211> 1093 <212> DNA <213> Artificial Sequence <220> <223> BIP2 SNP rs2306359 <220> <221> misc_feature <222> (159) <223> K represents T or G <400> 2 ccccggagga agccttggcc ccctcgtcct cttcgcccct ccaggccggc gacgtggggc 60 tgacggccag gtcgcaaaaa gcagggccga gcggagcccg ctcccctcgg tcggcggtgg 120 agaccccggc ccaatccccc ggccgcagcg gtacggcgkc ggcggcagca gctgacccgg 180 acrcagtgag aagccccggc ggaagtgata acatcccgac ctcctccggg cgcggcagcc 240 ggcggtggag gcggcggrgg cgtggcggcg gcggaggcgt ggcgtcttct tgaaacccac 300 acagcatccc cccaaccccc tcagacgtca cgggctagcg ctcgcgcctg cactcgcggg 360 gtcggccttc cttgcctgtg ggcggayccg tcgcgtcgac tccttattgg tggcgttact 420 acgcctcaag gcgtttcttc cctctttgct gtccgcaggg acgctgcctg gtagtttcag 480 ttacttttgt tttctttctg cctgccttta ttgatcatta cagtaggtga aattatcgat 540 atcaatata atattgtaat acaataggtg caattatcaa tatcaatatt attacaacag 600 gtgcaattat cagtatcatt acaataggtg caattatcag cgatcaataa tngtgaatta 660 taaaatggat gaaataaggg gaggaggtcg ggcgcggtgg tttacgcttg taatcccagc 720 actttgggag ggcgaggcgg gtggattacc ggaggtcaag agctcgagac cagcctggcc 780 aacatggtga cacctcgtct ctactaaaca aacaaaaatt agctgggcgt ggtggcgcgc 840 gtctgtaatc ccaactactc aggaggcgag ggcggaggcg gaggcggagt tggagtttgc 900 agtgagccga gatcgtgcca ttgcactcca gcttgggcaa cagagcaaaa actccgtctc 960 aaaaaaagg aaaaaaaaaaaaaaaaaaaaaaaaaaaaaaaaaaagaaataaggcga 1020 ggaaaaatat ccgaggcatc attttgggga gttaggcctc agaagcgacc ttagagtttg 1080 gtaatattkg agt 1093

Claims (8)

NCBI refSNP ID of the BNIP2 gene: rs754641 Composition for predicting the rejection reaction after organ transplantation comprising a probe or primer capable of detecting single nucleotide polymorphism as an active ingredient.
2. The composition of claim 1, wherein the single nucleotide polymorphism further comprises NCBI refSNP: rs2306359.
2. The composition of claim 1, wherein the organ transplant is a kidney transplant.
A method for detecting single nucleotide polymorphisms of the NCBI refSNP ID: rs754641 BNIP2 gene by analyzing the nucleotide sequence from a patient sample to provide information necessary for prediction of rejection after organ transplantation.
5. The method of claim 4, wherein the single nucleotide polymorphism further comprises NCBI refSNP: rs2306359.
. Methods for single nucleotide polymorphism analysis of the BNIP2 gene family for predicting rejection response after organ transplant including the following steps:
(a) isolating genomic DNA from an individual;
(b) determining the nucleotide sequence of the NCBI refSNP ID: rs754641 and rs2306359 sites;
(c) detecting a nucleotide sequence polymorphism in which G is replaced by A in the case of rs754641 and G is replaced by G in the case of rs2306359;
(d) determining a haplotype with the detected base sequence polymorphism; And

(e) Haplotypes H1 and H2 are high risk for rejection after organ transplantation, and H3 is low risk for rejection after organ transplantation.
A kit for predicting a rejection reaction after organ transplantation comprising a screening primer or a genotyping primer of a nucleotide sequence polymorphism selected from the group consisting of NCBI refSNP ID of rNBP2 gene: rs754641 and rs2306359.
8. The method of claim 7, wherein the single nucleotide polymorphism further comprises NCBI refSNP: rs2306359.

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