KR101894018B1 - Composition, microarray, and kits for predicting risk of periodontal disease, and method using the same - Google Patents

Abstract

The present invention provides a composition, a microarray, a kit according to one aspect, a method for predicting the risk of periodontal disease of an individual using the kit, and a method for analyzing SNP of an individual to provide information necessary for predicting the risk of periodontal disease. According to this, the risk of periodontal disease of an individual can be determined accurately and easily.

Description

TECHNICAL FIELD [0001] The present invention relates to a composition, a microarray, and a kit for predicting the risk of periodontal disease, and a method using the composition, a microarray, and a kit for predicting the risk of periodontal disease,

Kits, and microarrays for predicting the risk of periodontal disease through genetic analysis of single nucleotide polymorphisms of the IL-4, TENM2, or GPR141 gene, and methods of using the same.

Periodontal disease is a disease in which germs, such as Porphyromonas gingivalis, attack the gaps between the gums and teeth and damage the periodontal ligament and adjacent tissues. Periodontal disease is affected by a variety of causes, including biofilm, calculus, malnutrition, endocrine disfunction, systemic disease, smoking, and stress. According to the Health Insurance Statistical Yearbook of 2015, 13.33 million of the Korean health insurance subscribers received treatment for periodontal disease in 2015, and it is the second most common outbreak of total outpatient diagnosis. Recent studies have suggested that the importance of pre-emptying and preventing periodontal disease is emphasized, as the association between periodontal disease and genetic factors is known (Hong KW et al., 42 (8): 703-710, 2015). By utilizing genetic differences between people without periodontal disease and those with periodontal disease, people who are genetically at risk for periodontal disease can be used to prepare for disease in advance.

Single nucleotide polymorphism (SNP) is the most common form of genetic polymorphism on the human genome, and refers to a change in a particular nucleotide sequence on the genome. Genetically, SNPs can cause large differences in each individual depending on their location. For example, when a SNP is present in a position encoding a protein, it may affect the structure of the protein, thereby changing the function of the protein, and may also be related to disease. When the SNP is present in a non-coding region that does not encode a protein, that is, when it is present in a promoter or an intron, there is a difference in the expression level of the protein with respect to each of the SNPs, Abnormal protein may be expressed through alternative splicing.

Therefore, there is a need to identify SNPs that can predict or measure the risk of periodontal disease and to determine the risk of periodontal disease with high accuracy and sensitivity.

A composition for predicting the risk of periodontal disease, a microarray, or a kit.

A method for predicting the risk of periodontal disease of an individual or a method for providing information necessary for predicting the risk of periodontal disease.

One aspect is a polynucleotide consisting of a nucleic acid sequence of SEQ ID NO: 1, a first polynucleotide which is identical or complementary to 5 to 100 contiguous nucleic acid sequences comprising a polymorphic site of the 201 < th >

A second polynucleotide that is the same as or complementary to a 5-100 contiguous nucleic acid sequence comprising a polymorphic site of the 201 < th > nucleotide from its 5 ' -end in a polynucleotide consisting of the nucleic acid sequence of SEQ ID NO: 2; And

A polynucleotide consisting of a nucleotide sequence of SEQ ID NO: 3 and a third polynucleotide that is the same as or complementary to 5 to 100 contiguous nucleic acid sequences comprising a polymorphic site of the 201 < th > A composition for predicting the risk of periodontal disease including nucleotides is provided.

The polymorphic site refers to a site showing a single base polymorphism in the nucleic acid sequence. The term " single nucleotide polymorphism " (SNP) refers to a genetic change or mutation that exhibits a single nucleotide difference in a nucleic acid sequence. It may be a position where two or more contiguous nucleotide sequences are present in the population in a frequency of 1% or more or 5% or more.

The first polynucleotide, the second polynucleotide, and the third polynucleotide each comprise a polynucleotide consisting of the nucleotide sequence of SEQ ID NO: 1, a polynucleotide consisting of the nucleotide sequence of SEQ ID NO: 2, 10 to 100, 10 to 80, 10 to 60, 10 to 40, 10 to 10, 10, 20, 30, 40, 50 or more nucleotides comprising polymorphic sites of the 201 & To 20 contiguous nucleic acid sequences.

The first polynucleotide, the second polynucleotide, the third polynucleotide, or a combination thereof may be a primer or a probe.

The term "primer" refers to a single-stranded polynucleotide that can act as a starting point in the polymerization of a nucleotide by a polymerase. For example, the primer can serve as a starting point for template-directed DNA synthesis in the presence of suitable conditions, i.e., four different nucleoside triphosphates and polymerases, at the appropriate temperature and in the appropriate buffer Lt; RTI ID = 0.0 > polynucleotides < / RTI > The appropriate length of the primer may vary depending on a variety of factors, such as temperature and use of the primer. The primer may be 15 to 30 nucleotides in length. For example, the shorter the length of the primer, the more stable the hybridization complex with the template at low annealing temperatures. The length of the primer is about 5 to about 100 nucleotides (hereinafter referred to as 'nt'), about 10 to about 80 nt, about 10 to about 60 nt, about 10 to about 50 nt, about 10 to about 40 nt, 10 to about 30 nt, or about 10 to about 20 nt.

The term "probe" refers to an oligonucleotide capable of specifically binding to a target nucleic acid. The probe may be bound to a labeling substance. The probe may be from about 50 nt to about 400 nt in length, from about 100 nt to about 350 nt, from about 150 nt to about 300 nt, or from about 200 nt to about 250 nt in length.

The first polynucleotide may be a polynucleotide for analyzing a genotype of a single base polymorphism of interleukin-4 (IL-4) gene. Interleukin-4 is a cytokine that induces the differentiation of non-contact helper T cells into Th2 cells, stimulating the proliferation of activated B cells and T cells, and differentiating B cells into plasma cells. The IL-4 protein is expressed in human UniProt No. 1. Lt; RTI ID = 0.0 > P05112. ≪ / RTI > The first polynucleotide may be a polynucleotide for detecting the genotype of SNP ID rs2243250. The rs2243250 may be a polymorphic site of the 201 < th > nucleotide from its 5 ' -end at a polynucleotide consisting of the nucleic acid sequence of SEQ ID NO: 1. The rs2243250 may have a genotype of TT, CT, or CC. If the genotype of rs2243250 is CC, the risk of periodontal disease can be expected to increase.

The first polynucleotide may be a polynucleotide that is the same as or a complementary polynucleotide of 5-25 consecutive nucleic acid sequences comprising a polymorphic site of the 26th nucleotide from its 5'-terminus in a polynucleotide consisting of the nucleic acid sequence of SEQ ID NO: 3 have.

The second polynucleotide may be a polynucleotide for analyzing the genotype of a single nucleotide polymorphism of the teneurin transmembrane protein 2 (TENM2) gene. The TENM2 protein is expressed in human UniProt No. 1. Or a protein consisting of the amino acid sequence of Q9NT68. The second polynucleotide may be a polynucleotide for detecting the genotype of SNP ID rs4242220. The rs4242220 may be a polymorphic site of the 201 < th > nucleotide from its 5 ' -end at a polynucleotide consisting of the nucleic acid sequence of SEQ ID NO: 2. The rs4242220 may have a genotype of TT, GT, or GG. If the genotype of rs4242220 is GG, the risk of periodontal disease can be expected to increase.

The second polynucleotide may be a polynucleotide that is the same as or complementary to the 5 to 25 consecutive nucleic acid sequences comprising the polymorphic site of the 26th nucleotide from its 5'-end in a polynucleotide consisting of the nucleic acid sequence of SEQ ID NO: 4 have.

The third polynucleotide may be a polynucleotide for analyzing a genotype of a single base polymorphism of a G protein-coupled receptor 141 (GPR141) gene. The GPR141 protein is expressed in UniProt No. 1 in human. Or a protein consisting of the amino acid sequence of Q7Z602. The third polynucleotide may be a polynucleotide for detecting the genotype of SNP ID rs2392510. The rs2392510 may be a polymorphic site of the 201 < th > nucleotide from its 5 ' -end at a polynucleotide consisting of the nucleic acid sequence of SEQ ID NO: 2. The rs2392510 may have CC, CT, or TT genotypes. If the genotype of rs2392510 is TT, the risk of periodontal disease can be expected to increase.

Wherein the third polynucleotide comprises a polynucleotide consisting of the same or a complementary nucleic acid sequence as the 5 to 25 consecutive nucleic acid sequences comprising a polymorphic site of the 26th nucleotide from its 5'-end in a polynucleotide consisting of the nucleic acid sequence of SEQ ID NO: 5 0.0 > polynucleotides < / RTI >

The term "periodontal disease" refers to inflammation of the gingiva (gums), periodontal ligament, and bone tissue around the teeth. Periodontal disease can also be called taste. Periodontal disease includes gingivitis and periodontitis.

Another aspect is a polynucleotide which is identical or complementary to 5 to 100 contiguous nucleic acid sequences comprising a polymorphic site of the 201 < th > nucleotide from its 5 ' -end in a polynucleotide consisting of the nucleic acid sequence of SEQ ID NO: 1;

A polynucleotide which is the same as or complementary to 5 to 100 contiguous nucleic acid sequences comprising a polymorphic site of the 201 < th > nucleotide from its 5 ' -end in a polynucleotide consisting of the nucleic acid sequence of SEQ ID NO: 2; And

A polynucleotide selected from the group consisting of a polynucleotide consisting of the nucleotide sequence of SEQ ID NO: 3 and the same or complementary to 5 to 100 consecutive nucleotide sequences comprising the polymorphic site of the 201 < th > A microarray for predicting the risk of periodontal disease is provided.

The microarray may be a microarray for predicting the risk of periodontal disease for genotyping a single nucleotide polymorphism of the IL-4 gene, the TENM2 gene, the GPR141 gene, or a combination thereof.

A microarray refers to a polynucleotide immobilized at a high density in a divided region of a substrate surface. The microarray may be arranged on the substrate at a density of, for example, 400 cells / cm 2 or more, 10 ³ cells / cm 2 or 10 ⁴ cells / cm 2 or more.

The polynucleotide immobilized on the microarray may be DNA or RNA. The polynucleotide may be single-stranded or double-stranded. The polynucleotide can include a natural nucleotide, an analog of a natural nucleotide, a nucleotide in which the sugar, base or phosphate site of a natural nucleotide is modified, a peptide nucleic acid (PNA), and combinations thereof. The polynucleotide may be a detectable label (e.g., Cy3 or Cy5 fluorescent material) attached at its 3'-end, intermediate, or 5'-end.

Wherein the polynucleotide immobilized on the microarray has a length of about 5 to about 100 nt, about 10 to about 80 nt, about 10 to about 60 nt, about 10 to about 50 nt, about 10 to about 40 nt, 30 nt, or from about 10 to about 20 nt.

Another aspect is a polynucleotide which is identical or complementary to 5 to 100 contiguous nucleic acid sequences comprising a polymorphic site of the 201 < th > nucleotide from its 5 ' -end in a polynucleotide consisting of the nucleic acid sequence of SEQ ID NO: 1;

A polynucleotide which is the same as or complementary to 5 to 100 contiguous nucleic acid sequences comprising a polymorphic site of the 201 < th > nucleotide from its 5 ' -end in a polynucleotide consisting of the nucleic acid sequence of SEQ ID NO: 2; And

A polynucleotide selected from the group consisting of a polynucleotide consisting of the nucleotide sequence of SEQ ID NO: 3 and the same or complementary to 5 to 100 consecutive nucleotide sequences comprising the polymorphic site of the 201 < th > A kit for predicting the risk of periodontal disease is provided.

The kit may be a kit for predicting the risk of periodontal disease for genotyping a single nucleotide polymorphism of the IL-4 gene, the TENM2 gene, the GPR141 gene, or a combination thereof.

The kit may further include reagents necessary for the polymerization reaction, such as dNTP, polymerase, and coloring agent.

Another aspect is a method for analyzing a genotype of a polymorphic site of a polynucleotide selected from the group consisting of SEQ ID NOS: 1 to 3 from a nucleic acid sample obtained from an individual,

The polymorphic site is the 201 < th > nucleotide from the 5 ' -terminal in SEQ ID NO: 1,

The 201 th nucleotide from the 5'-end in SEQ ID NO: 2, or

Is the 201 th nucleotide from the 5'-end in SEQ ID NO: 3; And

And calculating the risk of periodontal disease of the individual from the analyzed genotype.

The method may further comprise obtaining a nucleic acid sample from the biological sample of the individual. The subject may be a mammal, including a human. The biological sample refers to a sample obtained from an organism. The biological sample may be, for example, tissue, urine, mucus, saliva, tears, blood, plasma, serum, sputum, spinal fluid, pleural effusion, aspiration nipple, lymphatic fluid, airway fluid, intestinal fluid, , Cerebrospinal fluid, intracranial fluid, ascites, cystic tumor fluid, positive fluid, or a combination thereof. The step of obtaining the nucleic acid sample from the biological sample may be performed by a conventional DNA separation method. For example, the target nucleic acid can be amplified by polymerase chain reaction (LCR), ligase chain reaction (LCR), transcription amplification, or real-time nucleic acid sequence based amplification (NASBA) and purified.

The method comprises analyzing a genotype of a polymorphic site of a polynucleotide selected from the group consisting of SEQ ID NOS: 1 to 3 from a nucleic acid sample obtained from an individual.

The polymorphic site may be the 201 < th > nucleotide from the 5 ' -terminal in SEQ ID NO: The polymorphic site may be the 201 < th > nucleotide from the 5 ' -terminal in SEQ ID NO: 2. The polymorphic site may be the 201 < th > nucleotide from the 5 ' -terminal in SEQ ID NO: 3.

Wherein the step of analyzing the genotype comprises: a primer comprising a nucleic acid sequence identical or complementary to a nucleic acid sequence comprising a polymorphic site of the 201 < th > nucleotide from its 5 ' -terminal in a polynucleotide consisting of the nucleic acid sequence of SEQ ID NO: 1;

A primer comprising a nucleic acid sequence identical or complementary to a nucleic acid sequence comprising a polymorphic site of the 201 < th > nucleotide from its 5 ' -end in a polynucleotide consisting of the nucleic acid sequence of SEQ ID NO: 2; And

A primer comprising a nucleic acid sequence identical or complementary to a nucleic acid sequence comprising a polymorphic site of a 201 < th > nucleotide from its 5 ' -terminal in a polynucleotide consisting of the nucleic acid sequence of SEQ ID NO: 3 .

The step of analyzing the genotype may be performed using the first polynucleotide, the second polynucleotide, the third polynucleotide, or a combination thereof. The step of analyzing the genotype may be polymerase chain reaction (PCR), molecular beacon, primer extension, and the like.

Wherein the step of analyzing the genotype comprises: hybridizing the nucleic acid sample to a microarray to which the first polynucleotide, the second polynucleotide, the third polynucleotide, or a combination thereof is immobilized; And analyzing the hybridization results. The hybridization of nucleic acids on a microarray and the detection of hybridization results are well known in the art. The detection can be accomplished, for example, by labeling the nucleic acid sample with a labeling substance capable of generating a detectable signal comprising a fluorescent material, such as Cy3 and Cy5, and then hybridizing on the microarray and generating The hybridization result can be detected.

And a step of assigning a genotype score to the genotype of the polymorphic site.

The polymorphic site of the polynucleotide of SEQ ID NO: 1 can be assigned values of alpha ₁ , alpha ₂ , and alpha ₃ to the genotypes TT, CT, and CC, respectively. The values of? ₁ ,? ₂ , and? ₃ may be an integer of 0 to 2. For example, the score for genotype TT is 0, the score for genotype CT is 1, and the score for genotype CC is 2.

The polymorphic site of the polynucleotide of SEQ ID NO: 2 can be assigned values of beta ₁ , beta ₂ , and beta ₃ to the genotypes TT, GT, and GG, respectively. The values of? ₁ ,? ₂ , and? ₃ may be an integer of 0 to 2. For example, the score for genotype TT is 0, the score for genotype GT is 1, and the score for genotype GG is 2.

The polymorphic sites of the polynucleotide of SEQ ID NO: 3 can be assigned values of? ₁ ,? ₂ , and? ₃ to the genotypes CC, CT, and TT, respectively. The values of? ₁ ,? ₂ , and? ₃ may be an integer of 0 to 2. For example, the score for genotype CC is 0, the score for genotype CT is 1, and the score for genotype TT is 2.

The method includes calculating the periodontal disease risk of an individual from the analyzed genotype.

The step of calculating the periodontal disease risk of an individual can calculate the periodontal disease risk according to the following equation:

Periodontal disease risk = b + Σ (genotype score * a).

In the above equation, the genotypic score is a score assigned to a genotype of a polymorphic site of a polynucleotide analyzed in an individual.

The genotype of the polymorphism of the polynucleotide of SEQ ID NO: 1 may be any one of the values of? ₁ ,? ₂ , and? ₃ . α ₁ , α ₂ , and α ₃ may be the values assigned to the genotypes TT, CT, and CC for the polymorphic site of the polynucleotide of SEQ ID NO: 1. For example, when the genotype of the individual for SEQ ID NO: 1 is CC, the genotype score is? ₃ .

May be any of the values of? ₁ ,? ₂ , and? ₃ depending on the genotype of the polymorphism of the polynucleotide of SEQ ID NO: 2. β ₁ , β ₂ , and β ₃ may be the values assigned to the genotypes TT, GT, and GG for the polymorphic site of the polynucleotide of SEQ ID NO: 2. For example, if the genotype for the individual SEQ ID NO: 2 is GT, the genotype score is beta ₂ .

May be any one of the values of? ₁ ,? ₂ , and? ₃ according to the genotype of the polymorphism of the polynucleotide of SEQ ID NO: 3. γ ₁ , γ ₂ , and γ ₃ may be the values assigned to the genotypes CC, CT, and TT for the polymorphic site of the polynucleotide of SEQ ID NO: 3. For example, a case where the genotype for the SEQ ID NO: 3 of the object CC, the genotype score γ _1.

A may be a constant calculated by analyzing the genotype between the periodontal disease patient group and the control group without periodontal disease in the logistic regression model. The a may be an effect size value calculated in a logistic regression analysis model.

The a may be a value of 0.1 to 0.5, 0.12 to 0.45, 0.14 to 0.4, 0.16 to 0.35, 0.18 to 0.3, 0.2 to 0.28, 0.21 to 0.26, or 0.22 to 0.24 for the polymorphic genotype of the polynucleotide of SEQ ID NO: have. For example, for a genotype of SEQ ID NO: 1, the a may be 0.233.

Wherein a is a polynucleotide polymorphism of the polynucleotide of SEQ ID NO: 2 in the range of 0.7 to 1.2, 0.72 to 0.18, 0.74 to 1.16, 0.76 to 1.14, 0.78 to 1.12, 0.80 to 1.1, 0.82 to 1.08, 0.84 to 1.06, 0.86 to 1.04 , 0.88 to 1.02, 0.9 to 1, 0.9 to 0.98, 0.9 to 0.96, 0.9 to 0.94, or 0.9 to 0.92. For example, for a genotype of SEQ ID NO: 2, the a may be 0.912.

A is a polymorphism of the polynucleotide polymorphism of SEQ ID NO: 3 in the range of 0.8 to 1.3, 0.82 to 1.28, 0.84 to 1.26, 0.86 to 1.24, 0.88 to 1.22, 0.9 to 1.2, 0.92 to 1.18, 0.94 to 1.16, 0.96 to 1.14 , 0.98 to 1.12, 1 to 1.1, 1 to 1.08, 1 to 1.06, 1 to 1.04, or 1 to 1.02. For example, for a genotype of SEQ ID NO: 3, the a may be 1.014.

B can be a constant calculated by analyzing the genotype between the periodontal disease patient group and the control group without periodontal disease in the logistic regression model. And b may be a rare-coefficient (or y-intercept) value calculated in a logistic regression analysis model. The value b is a value of -0.5 to -1.5, -0.55 to -1.4, -0.6 to -1.3, -0.65 to -1.2, -0.7 to -1.1, -0.75 to -1, or -0.8 to -0.9. For example, b may be -0.814.

The method may further comprise the step of determining the risk of periodontal disease of the subject to any one of good, attention, attention, and concentrated management according to the calculated periodontal disease risk. If the calculated risk of periodontal disease is less than 1, the risk of periodontal disease of the individual can be determined as a good grade. If the calculated risk of periodontal disease is greater than 1 and less than 2, the risk of periodontal disease in the individual can be determined as the degree of concern. If the calculated risk of periodontal disease is more than 2 and less than 3, the risk of periodontal disease in the individual can be determined by the attentive grade. If the risk of periodontal disease is more than 3, the risk of periodontal disease of the individual can be determined as the concentration level.

According to a method of analyzing SNP of an individual in order to provide information necessary for predicting the risk of periodontal disease or predicting the risk of periodontal disease using primers, microarrays, kits, and the method of the present invention, The risk of periodontal disease can be determined accurately and easily.

Fig. 1 is a graph showing the ratio (%) of the number of individuals to the genotype score in the patient group and the control group.

Hereinafter, the present invention will be described in more detail with reference to Examples. However, these embodiments are for illustrative purposes only, and the scope of the present invention is not limited to these embodiments.

Example  One. Periodontal disease  Risk assessment

1. Preparation of nucleic acid samples

Blood of each subject was obtained from the patients diagnosed with periodontal disease (48 patients, patients over 40 years old) and the control group (23 patients, under 60 years old) who were diagnosed as not suffering from periodontal disease (Daegu Mir- ). A group without periodontal disease up to age 60 is genetically more resistant to periodontal disease. In addition, the group with periodontal disease from age 40 belongs to the genetically high defense group against periodontal disease.

Genomic DNA was isolated from the obtained blood of the subject using QIAamp DNA Blood Mini Kit (QIAGEN). The isolated DNA was stored at -20 ° C until use.

2. Detection of single nucleotide polymorphisms

Single nucleotide polymorphism (SNP) of IL-4, TENM2, or GPR141 was detected using the TaqMan probe method using the prepared genomic DNA as a template.

Specifically, 20 ng of genomic DNA prepared as described in 1., 12.5 μl of 2 × TaqMan Universal PCR master mix, 20 × primer of TaqMan SNP Genotyping Assay specific to the SNP of the target gene, and 1.25 μl TaqMan probe dye mix, And 11.25 占 퐇 of DNase free sterile water were mixed to prepare a mixture of 25 占 퐇 of the final volume. Target sequences including target genes and SNPs are shown in Table 1 below.

Target gene Reference SNP
(NCBI number)  Analytics ID Target sequence IL-4 rs2243250
(SEQ ID NO: 1) C__16176216_10 5'-ACACCTAAACTTGGGAGAACATTGT [C / T] CCCCAGTGCTGGGGTAGGAGAGTCT-3 '(SEQ ID NO: 4) TENM2 rs4242220
(SEQ ID NO: 2) C__27106157_10 5'-AAATGTCCTGTAATGGCCATTTCTG [G / T] GTCGGTTTCCTGTTCCTTTCCCTAA-3 '(SEQ ID NO: 5) GPR141 rs2392510
(SEQ ID NO: 3) C__11835423_10 5'-AGAGCAACCACCCACCCAGCTGGTA [C / T] GTTTTCTTCTCTTCCACGCCCACTG-3 '(SEQ ID NO: 6)

The nucleic acid in "[]" in the target sequence of Table 1 represents the SNP of the target gene.

The prepared mixture was dispensed into 96-well plates and subjected to polymerase chain reaction (PCR). The PCR conditions were 95 ° C for 10 minutes, 95 ° C for 15 seconds, 60 ° C for 1 minute, and 40 cycles were repeated. Then, the SNP of the target gene was detected using a real-time PCR system (TaqMan Mastermix, ABI).

3. Periodontal disease risk assessment

Genotypes of target genes were analyzed by detecting SNP in periodontal disease patients and controls. SNP genotypes were assigned a score of 0, 1, or 2, and SNP genotypic results between periodontal disease patients and controls were analyzed by logistic regression. From the logistic regression analysis, β value was used as the effect size. The genotype and calculated magnitude of influence are shown in Table 2 below.

Target gene Reference SNP Influence size Genotype score 0 One 2 Y Intercept -0.814 IL-4 rs2243250 0.233 TT CT CC TENM2 rs4242220 0.912 TT GT GG GPR141 rs2392510 1.014 CC CT TT

The score of the genetic type (SG) was calculated from the influence magnitude of the above Table 2 according to the following equation (1).

[Formula 1]

SG = -0.814 + [(genotype score of IL-4 * 0.233) + (genotype score of TENM2 * 0.912) + (genotype score of GPR141 * 1.014)]

The genotype scores were calculated using the formula 1 in the periodontal disease patients and the control group, and the calculated genotype score and periodontal disease risk are shown in Table 3 below.

Periodontal disease risk Good Attention caution Concentration management Genotype score (SG) <1 1 < = < 2 2 < = < 3 3 <= Control group (persons) 23 14 9 0 0 Patient group (persons) 48 17 23 7 One

As shown in Table 3, the genotype score of the periodontal disease patient group was significantly higher than the genotype score of the control group. Thus, it can be seen that information on the risk of periodontal disease can be more accurately provided to an individual by discriminating the risk of periodontal disease using a logistic regression model of a specific gene SNP.

<110> Theragen Etex Co., Ltd. <120> Composition, microarray, and kits for predicting risk of          periodontal disease, and method using the same <130> PN117015KR <160> 6 <170> Kopatentin 2.0 <210> 1 <211> 401 <212> DNA <213> Artificial Sequence <220> <223> rs2243250 <220> <221> variation <201> <223> single nucleotide polymorphism [C / T] <400> 1 tgcatttcag cctgggtgac agagtgagac cctgtctcag aaaaaaaaaa aaaaaagtca 60 ttcctgaaac ctcagaatag acctaccttg ccaagggctt ccttatgggt aaggacctta 120 tggacctgct gggacccaaa ctaggcctca cctgatacga cctgtccttc tcaaaacacc 180 taaacttggg agaacattgt cccccagtgc tggggtagga gagtctgcct gttattctgc 240 ctctatgcag agaaggagcc ccagatcagc ttttccatga caggacagtt tccaagatgc 300 cacctgtact tggaagaagc caggttaaaa tacttttcaa gtaaaacttt cttgatatta 360 ctctatcttt ccccaggagg actgcattac aacaaattcg g 401 <210> 2 <211> 401 <212> DNA <213> Artificial Sequence <220> <223> rs4242220 <220> <221> variation <201> <223> single nucleotide polymorphism [G / T] <400> 2 tgtgttgtgc tcaacggaaa ttttccacat ctccatctct gtaactccac tgtcaagagc 60 aatctcttag gcactgcctg ccattgtgat tattgccaat atcactgaat taaggtaatc 120 aggccacatt catcgcatgc acttggagca gaagctgaaa ccacaggtaa aattgaaatg 180 tcctgtaatg gccatttctg ggtcggtttc ctgttccttt ccctaaagtg catttgcttt 240 tgcttttcct ccttgcctgc tgagatcatt gtgtgggtgt gagtgtctga gggttcaggt 300 tgataagttg caaagggaag aggggataaa aagccacaca gcttgaataa ataaatgcat 360 gaaaatatgt ggtactctag catgccctac cgccatgtct c 401 <210> 3 <211> 401 <212> DNA <213> Artificial Sequence <220> <223> rs2392510 <220> <221> variation <201> <223> single nucleotide polymorphism [C / T] <400> 3 cctctctata tgcaagtgac agcgtgaatt ttgtaggacg ctttttcaca tacattttct 60 tttcatataa atcgtaatga ttgaccagag acccttgcaa atagcctggg catggacaaa 120 tcctgagagc 180 aaccacccac ccagctggta cgttttcttc tcttccacgc ccactgcctc ttcctctttc 240 cctttccctc cagagcttcc ctgaccctct tctcctcccc tttacctcct tcctgttaaa 300 aagtgtctgg tggtggagcc agacagaact gtagtcagag ccttgtggga tttctcaggg 360 gcagggtaac ctcaggtcct gaacctccca gaacgtctac a 401 <210> 4 <211> 51 <212> DNA <213> Artificial Sequence <220> <223> Target sequence for rs2243250 <220> <221> variation (26) <223> single nucleotide polymorphism [C / T] <400> 4 acacctaaac ttgggagaac attgtccccc agtgctgggg taggagagtc t 51 <210> 5 <211> 51 <212> DNA <213> Artificial Sequence <220> <223> Target sequence for rs4242220 <220> <221> variation (26) <223> single nucleotide polymorphism [G / T] <400> 5 aaatgtcctg taatggccat ttctgggtcg gtttcctgtt cctttcccta a 51 <210> 6 <211> 51 <212> DNA <213> Artificial Sequence <220> <223> Target sequence for rs2392510 <220> <221> variation (26) <223> single nucleotide polymorphism [C / T] <400> 6 agagcaacca cccacccagc tggtacgttt tcttctcttc cacgcccact g 51

Claims (16)

A first polynucleotide which is identical or complementary to 5 to 100 contiguous nucleic acid sequences comprising a polymorphic site of the 201 &lt; th &gt; nucleotide from its 5 ' -terminal at a polynucleotide consisting of the nucleic acid sequence of SEQ ID NO: 1;
A second polynucleotide that is the same as or complementary to a 5-100 contiguous nucleic acid sequence comprising a polymorphic site of the 201 &lt; th &gt; nucleotide from its 5 &apos; -end in a polynucleotide consisting of the nucleic acid sequence of SEQ ID NO: 2; And
A polynucleotide consisting of a nucleic acid sequence of SEQ ID NO: 3 and a polynucleotide sequence of 5'-100 nucleotides from the 5'-terminus thereof, and a third polynucleotide which is identical or complementary to 5-100 consecutive nucleotide sequences Compositions for predicting risk. The method according to claim 1,
A first polynucleotide which is the same as or complementary to the 5 to 25 consecutive nucleic acid sequences comprising the polymorphic site of the 26th nucleotide from the 5'-end thereof in a polynucleotide consisting of the nucleic acid sequence of SEQ ID NO: 4;
A second polynucleotide which is the same as or complementary to the 5 to 25 consecutive nucleic acid sequences comprising the polymorphic site of the 26th nucleotide from the 5'-end thereof in a polynucleotide consisting of the nucleic acid sequence of SEQ ID NO: 5; And
A polynucleotide consisting of a nucleotide sequence of SEQ ID NO: 6, and a third polynucleotide that is identical or complementary to 5 to 25 contiguous nucleic acid sequences comprising a polymorphic site of the 26 &lt; th &gt; 0.0 &gt; nucleotides. &Lt; / RTI &gt; A first polynucleotide which is identical or complementary to 5 to 100 contiguous nucleic acid sequences comprising a polymorphic site of the 201 &lt; th &gt; nucleotide from its 5 ' -terminal at a polynucleotide consisting of the nucleic acid sequence of SEQ ID NO: 1;
A second polynucleotide that is the same as or complementary to a 5-100 contiguous nucleic acid sequence comprising a polymorphic site of the 201 &lt; th &gt; nucleotide from its 5 &apos; -end in a polynucleotide consisting of the nucleic acid sequence of SEQ ID NO: 2; And
A polynucleotide consisting of a nucleic acid sequence of SEQ ID NO: 3 and a polynucleotide sequence of 5'-100 nucleotides from the 5'-terminus thereof, and a third polynucleotide which is identical or complementary to 5-100 consecutive nucleotide sequences Microarray to predict risk. A first polynucleotide which is identical or complementary to 5 to 100 contiguous nucleic acid sequences comprising a polymorphic site of the 201 &lt; th &gt; nucleotide from its 5 ' -terminal at a polynucleotide consisting of the nucleic acid sequence of SEQ ID NO: 1;
A second polynucleotide that is the same as or complementary to a 5-100 contiguous nucleic acid sequence comprising a polymorphic site of the 201 &lt; th &gt; nucleotide from its 5 &apos; -end in a polynucleotide consisting of the nucleic acid sequence of SEQ ID NO: 2; And
A polynucleotide consisting of a nucleic acid sequence of SEQ ID NO: 3 and a polynucleotide sequence of 5'-100 nucleotides from the 5'-terminus thereof, and a third polynucleotide which is identical or complementary to 5-100 consecutive nucleotide sequences A kit for predicting risk. Analyzing a genotype of a polymorphic site of the polynucleotide of SEQ ID NOS: 1 to 3 from a nucleic acid sample obtained from an individual,
Wherein the polymorphic site is the 201 &lt; th &gt; nucleotide from the 5 ' -terminal in SEQ ID NO: 1,
The 201 th nucleotide from the 5'-end in SEQ ID NO: 2, and
Is the 201 th nucleotide from the 5'-end in SEQ ID NO: 3; And
Calculating the risk of periodontal disease of the individual according to the following equation from the analyzed genotype,
Periodontal disease risk = b + Σ (genotype score * a)
In the above equation,
Wherein the genotypic score is a score assigned to a genotype of a polymorphic site of the polynucleotide analyzed in the subject,
A genotype score of? ₁ ,? ₂ , and? ₃ assigned to the genotypes TT, CT, and CC at the polymorphic site of the polynucleotide of SEQ ID NO: 1,
A genotype score of? ₁ ,? ₂ , and? ₃ assigned to the genotypes TT, GT, and GG at the polymorphic site of the polynucleotide of SEQ ID NO: 2,
A genotype score of? ₁ ,? ₂ , and? ₃ assigned to the genotypes CC, CT, and TT at the polymorphic site of the polynucleotide of SEQ ID NO: 3,
Wherein a is a value of 0.1 to 0.5 for the polymorphism of the polynucleotide of SEQ ID NO: 1, 0.7 to 1.2 for the polymorphism of the polynucleotide of SEQ ID NO: 2, or a polynomial of polynucleotide of SEQ ID NO: 0.0 &gt; 1.3 &lt; / RTI &gt; And
And b is a value of -0.5 to -1.5. The method of claim 5, wherein analyzing the genotype comprises:
A primer comprising a nucleic acid sequence identical or complementary to a nucleic acid sequence comprising a polymorphic site of a 201 &lt; th &gt; nucleotide from its 5 ' -terminal at a polynucleotide consisting of the nucleic acid sequence of SEQ ID NO: 1;
A primer comprising a nucleic acid sequence identical or complementary to a nucleic acid sequence comprising a polymorphic site of the 201 &lt; th &gt; nucleotide from its 5 &apos; -end in a polynucleotide consisting of the nucleic acid sequence of SEQ ID NO: 2; And
A primer comprising a nucleic acid sequence identical or complementary to a nucleic acid sequence comprising a polymorphic site of a 201 &lt; th &gt; nucleotide from its 5 &apos; -terminal in a polynucleotide consisting of the nucleic acid sequence of SEQ ID NO: 3 How it is. The method of claim 5, wherein analyzing the genotype comprises:
A first polynucleotide that is identical or complementary to at least five consecutive nucleic acid sequences comprising a polymorphic site of the 201 &lt; th &gt; nucleotide from its 5 ' -terminal at a polynucleotide consisting of the nucleic acid sequence of SEQ ID NO: 1;
A second polynucleotide that is the same as or complementary to at least five consecutive nucleic acid sequences comprising a polymorphic site of the 201 &lt; th &gt; nucleotide from its 5 &apos; -end in a polynucleotide consisting of the nucleic acid sequence of SEQ ID NO: 2; And
A polynucleotide consisting of a nucleotide sequence of SEQ ID NO: 3 and a polynucleotide selected from the group consisting of a third polynucleotide identical or complementary to five or more consecutive nucleic acid sequences comprising a polymorphic site of the 201 &lt; th &gt; Hybridizing the nucleic acid sample to a fixed microarray; And
And analyzing the hybridization result. 6. The method of claim 5, further comprising genotyping a genotype of the polymorphic site. delete The method according to claim 5, wherein the genotyping score of? ₁ ,? ₂ , and? ₃ is an integer of 0 to 2. delete The method according to claim 5, wherein the genotypic score of? ₁ ,? ₂ , and? ₃ is an integer of 0 to 2. delete The method according to claim 5, wherein the genotype score of? ₁ ,? ₂ ,? ₃ is an integer from 0 to 2. delete The method according to claim 5, wherein the calculated periodontal disease risk is
1, the risk of periodontal disease of an individual is determined as a good grade,
1 or more and less than 2, the risk of periodontal disease of an individual is determined as a degree of concern,
2 or more and less than 3, the risk of periodontal disease of an individual is determined by the grade of caution, and
3 &gt; or more, the risk of periodontal disease of the subject is determined as the concentration level.

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