KR20100055367A - Polynucleotides comprising single nucleotide polymorphism, microarrays and diagnostic kits comprising the same, and detection methods using the same - Google Patents

Abstract

PURPOSE: A polynucleotide containing single-nucleotide polymorphism(SNP) which is derived from MTHFR, AUTS2, RELN, LAMB1, WNT2, FOXP2, and ST7 is provided to use as a clinical diagnosis marker for autism. CONSTITUTION: A nucleotide for clinically diagnosing autism comprises 10 or more serial DNA sequences contain 301th nucleotide T. The length of the polynucleotide is 10-100. A primer for amplifying diagnosis contains the polynucleotide or complementary polynucleotide. A microarray comprises a probe for diagnosing autism for Korean.

Description

Polynucleotides comprising single nucleotide polymorphism, microarrays and diagnostic kits comprising the same, and detection methods using the same}

The present invention relates to autism (autism), a polynucleotide comprising a polynucleotide or a complementary nucleotide thereof; An amplification primer for autism diagnosis or a probe for autism diagnosis consisting of the polynucleotide or its complementary nucleotide; A microarray including the probe; And a detection method using the polynucleotide or its complementary nucleotide, which is useful for providing information necessary for diagnosing autism.

Human chromosomes are all 23 pairs, consisting of 22 pairs of homologous chromosomes and 1 pair of sex chromosomes. The 23 pairs of chromosomes received 23 from the father and 23 from the mother, and have a total of 46 chromosomes. With the successful completion of the Human Genome Project, it was found that about 99.9% of the sequences have the same genetic sequence among all human individuals. That is, only about 0.1% of the nucleotide sequence shows differences between individuals. Therefore, it is estimated that the difference in nucleotide sequence of 0.1% is the cause of the genetic difference between individuals.

There are largely four types of genetic variation that exist in the genome. First, genotyping according to the difference in the number of repeating sequences classified into satellites, minisatellite, and microsatellite (or STR; short tandem repeats) according to the length of the repeating sequence. This is my brother. Second, there are genotypic variations such as LINE, SINE, or transposable elements that are scattered sporadically in various places in the genome by retrotransposons. Third, there is a modification that occurs when a specific genetic site or nucleotide sequence is inserted or deleted. Fourth, it is the most common mutation, which accounts for about 90% or more of the total human genetic variant, and is a mutation form in which one nucleotide sequence is substituted.Single nucleotide polymorphism (SNP) is the most widely used in recent genetic diversity studies. : There is a single base polymorphism). SNP refers to the location where two allelic sequences (biallele) exist at a frequency of 1% or more in the human population, and is the most common form among the genetic variants possessed by humans, where one SNP exists every about 300bp. Is estimated to be. The genetic meaning of these SNPs makes a big difference to each individual depending on their location. For example, if a single nucleotide polymorphism is present at a position encoding a protein, it may affect the structure of the protein, so that the protein function may be changed, and it may be associated with a disease. As another example, when a single nucleotide polymorphism exists on another gene that does not encode a protein, that is, when it is present in a promoter or intron, the overall activity of the protein may decrease due to a difference in the expression level of the protein for each. In addition, the protein may be abnormally expressed through a denatured intron removal process (alternative splicing).

Various assays are used to confirm the genotype of such a single base polymorphism. The two most representative traditional methods are Restriction Fragment Length Polymorphism (RFLP) using restriction enzymes, and SSCP using a structure in which single-stranded DNA is stabilized by weak intermolecular bonds within itself affects electrophoresis. There is a way. Recently, depending on the scale, the method is largely divided into two types. The first method is a small scale method used to investigate single base polymorphism, TaqMan ^TM probe method using allele-specific hybridization, dynamic allele specificity using melting temperature (Tm). There are Dynamic Allele-Specific Hybridization (DASH) method and pyrosequencing using polymerization reaction. Secondly, as a method to identify the genotype of a large amount of single-base polymorphism, microarray technology uses the principle of Allele-Specific Extension to integrate and plant probes on a small substrate, and hybridize with target DNAs. The genotype of single base polymorphism is confirmed by performing and extension.

On the other hand, autism, also called Autism Spectrum Disorder (ASD), is a pervasive developmental disorder (PDD). It is a developmental disability. Autism appears in various forms and varies in severity from individual to individual.Autistic disorder, Rett's disorder, childhood disintegrative disorder, Asperger's syndrome, and general developmental disorder with specific inability ( Pervasive Developmental Disorder Not Otherwise Specified; PDD-NOS). PDD-NOS refers to a disorder that is close to autism, but does not meet the diagnostic criteria of the American Psychiatric Association, Diagnostic and Statistical Manual of Mental Disorder IV (DSM-IV), and has three main symptoms of autism. This refers to the case where all of these do not appear but some do not appear, and is defined as atypical autism disorder in the International Classification of Disease (ICD) prepared by the WHO.

Most of autism is childhood autism, with a prevalence of 2-5 per 10,000, and up to 20-50 per 10,000 with general developmental disorders such as Asperger's syndrome, and the male to female sex ratio is about 4:1, which is more frequent in males than females. [Gillberg C, Wing L. Autism: not an extremely rare disorder. Acta Psychiatr Scand; 99:399-406 (1999); Fombonne E. The epidemiology of autism: a review. Psychol Med; 29:769-786 (1999)]. The exact cause of autism has not yet been reported, but genetic factors have been suggested in many studies [Kaminsky Z, Wang SC, Petronis A. Complex disease, gender and epigenetics. Ann Med; 38:530-544 (2006)]. Siblings of autistic patients have a prevalence of autism of 2-4%, which is about 10 times higher than that of no autistic patients in the family [Folstein S, Rutter M. Genetic influences and infantile autism. Nature;265:726-728 (1977); Folstein S, Rutter M. Infantile autism: a genetic study of 21 twin pairs. J Child Psychol Psychiatry; 18:297-321 (1977)]. Also, in the twin study, the concordance rate for identical twins with autism was 36% and 0% for fraternal twins, and 85% for identical twins and about 10% for fraternal twins when the symptoms were broadly viewed. There are bars [Bailey A, Le Couteur A, Gottesman I, Bolton P, Simonoff E, Yuzda E, et al. Autism as a strongly genetic disorder: evidence from a British twin study. Psychol Med; 25:63-77 (1995)]. Through this, autism proves that genetic factors are an important factor in its pathogenesis. Recently, it has been suggested that autism is complicated and various interactive locuses are implicated through family and twin relationships studies [Pickles A , Bolton P, Macdonald H, Bailey A, Le Couteur A, Sim CH, et al. Latent-class analysis of recurrence risks for complex phenotypes with selection and measurement error: a twin and family history study of autism. Am J Hum Genet; 57:717-726 (1995); Risch N, Spiker D, Lotspeich L, Nouri N, Hinds D, Hallmayer J, et al. A genomic screen of autism: evidence for a multilocus etiology. Am J Hum Genet; 65:493-507 (1999); Folstein SE, Rosen-Sheidley B. Genetics of autism: complex aetiology for a heterogeneous disorder. Nat Rev Genet; 2:943-955 (2001)].

In order to uncover the genetic factors of autism, studies on the association with autism through various association analyzes and genome-wide screening are actively being conducted, and according to these studies, 7 times out of 22 homologous chromosomes in humans. Chromosomes have been reported as chromosomes that are highly associated with autism [IMGSAC. A full genome screen for autism with evidence for linkage to a region on chromosome 7q. International Molecular Genetic Study of Autism Consortium. Hum Mol Genet; 7:571-578 (1998); IMGSAC. Further characterization of the autism susceptibility locus AUTS1 on chromosome 7q. Hum Mol Genet; 10:973-982 (2001); Maestrini E, Paul A, Monaco AP, Bailey A.Identifying autism susceptibility genes. Neuron; 28:19-24 (2000); Philippe A, Martinez M, Guilloud-Bataille M, Gillberg C, Rastam M, Sponheim E, et al. Genome-wide scan for autism susceptibility genes. ParisAutism Research International Sibpair Study. Hum Mol Genet; 8:805-812 (1999)].

Despite many genetic studies to date, researchers have different opinions on how autism is passed on to offspring or which genes are involved, suggesting the possibility of genetic heterogeneity or involvement of multiple genes. I did. In addition, there are still no physiological or biological diagnostic criteria for the diagnosis of autism. Therefore, there is a need in the art to find an objective molecular biological indicator capable of diagnosing and predicting autism and judging a detailed disease.

The present inventors selected specific genes, namely MTHFR, AUTS2, RELN, LAMB1, WNT2, FOXP2, and ST7 genes, as candidate genes among chromosome 7 reported as a chromosome having a lot of association with autism, and at the molecular biological level for the diagnosis of autism. Various studies have been conducted to develop diagnostic indicators. As a result, surprisingly, it was found that only some of the monobasic polymorphisms showed statistically significant differences between the autism patient group and the normal group, and the analyzed monobasic polymorphism was a molecular biological index for the diagnosis of autism. Found that it could function.

Accordingly, an object of the present invention is to provide a polynucleotide containing a single polymorphism related to autism or a complementary polynucleotide thereof.

In addition, an object of the present invention is to provide a primer for amplification for diagnosis of autism or a probe for autism diagnosis consisting of the polynucleotide or its complementary nucleotide.

In addition, an object of the present invention is to provide a microarray including the probe.

In addition, an object of the present invention is to provide a detection method using the polynucleotide or its complementary nucleotide, which is useful for providing information necessary for diagnosing autism.

According to an aspect of the present invention, in the polynucleotide consisting of the DNA sequence of SEQ ID NO: 2, the 301st base (polymorphic site) is C, and the polynucleotide consisting of 10 or more consecutive DNA sequences including the 301st base ( Hereinafter referred to as "polynucleotide (b)"); In the polynucleotide consisting of the DNA sequence of SEQ ID NO: 3, the 301st base (polymorphic site) is A, and the polynucleotide consisting of 10 or more consecutive DNA sequences including the 301st base (hereinafter, "polynucleotide (c) "Referred to as"); In the polynucleotide consisting of the DNA sequence of SEQ ID NO: 4, the 401th base (polymorphic site) is G, and the polynucleotide consisting of 10 or more consecutive DNA sequences including the 401th base (hereinafter, "polynucleotide (d)) "Referred to as"); In the polynucleotide consisting of the DNA sequence of SEQ ID NO: 5, the 201st base (polymorphic site) is A, and the polynucleotide consisting of 10 or more consecutive DNA sequences including the 201th base (hereinafter, "polynucleotide (e)) "Referred to as"); In the polynucleotide consisting of the DNA sequence of SEQ ID NO: 8, the 301st base (polymorphic site) is A, and the polynucleotide consisting of 10 or more consecutive DNA sequences including the 301st base (hereinafter referred to as "polynucleotide (h) "Referred to as"); In the polynucleotide consisting of the DNA sequence of SEQ ID NO: 9, the 301st base (polymorphic site) is A, and the polynucleotide consisting of 10 or more consecutive DNA sequences including the 301st base (hereinafter, "polynucleotide (i) "Referred to as"); In the polynucleotide consisting of the DNA sequence of SEQ ID NO: 10, the 301st base (polymorphic site) is C, and the polynucleotide consisting of 10 or more consecutive DNA sequences including the 301st base (hereinafter, "polynucleotide (j) "Referred to as"); In the polynucleotide consisting of the DNA sequence of SEQ ID NO: 13, the 201 th base (polymorphic site) is A, and the polynucleotide consisting of 10 or more consecutive DNA sequences including the 201 th base (hereinafter, "polynucleotide (m) "Referred to as"); In the polynucleotide consisting of the DNA sequence of SEQ ID NO: 15, the 301st base (polymorphic site) is T, and the polynucleotide consisting of 10 or more consecutive DNA sequences including the 301st base (hereinafter referred to as "polynucleotide (o) "Referred to as"); In the polynucleotide consisting of the DNA sequence of SEQ ID NO: 16, the 163th base (polymorphic site) is G, and the polynucleotide consisting of 10 or more consecutive DNA sequences including the 163th base (hereinafter, "polynucleotide (p) "Referred to as"); In the polynucleotide consisting of the DNA sequence of SEQ ID NO: 17, the 301st base (polymorphic site) is T, and the polynucleotide consisting of 10 or more consecutive DNA sequences including the 301st base (hereinafter, "polynucleotide (q) "Referred to as"); In the polynucleotide consisting of the DNA sequence of SEQ ID NO: 18, the 477th base (polymorphic site) is A, and the polynucleotide consisting of 10 or more consecutive DNA sequences including the 477th base (hereinafter, "polynucleotide (r) "Referred to as"); In the polynucleotide consisting of the DNA sequence of SEQ ID NO: 19, the 301st base (polymorphic site) is C, and the polynucleotide consisting of 10 or more consecutive DNA sequences including the 301st base (hereinafter, "polynucleotide (s) "Referred to as"); And in the polynucleotide consisting of the DNA sequence of SEQ ID NO: 20, the 249th base (polymorphic site) is C, and the polynucleotide consisting of 10 or more consecutive DNA sequences including the 249th base (hereinafter, "polynucleotide (t A polynucleotide selected from the group consisting of) or a complementary nucleotide thereof is provided.

In addition, according to another aspect of the present invention, a primer for amplification for diagnosis of autism, consisting of the polynucleotide or a complementary polynucleotide thereof, is provided.

Further, according to another aspect of the present invention, there is provided a probe for diagnosing autism and a microarray including the same, consisting of the polynucleotide or a complementary polynucleotide thereof.

In addition, according to another aspect of the present invention, there is provided a kit for diagnosing autism, comprising the polynucleotide or a complementary polynucleotide thereof.

In addition, according to another aspect of the present invention, in order to provide information necessary for diagnosing autism, (i) obtaining a nucleic acid sample from the specimen; And (ii) analyzing the nucleotide sequence of a polymorphic site of a polynucleotide selected from one or more polynucleotides of SEQ ID NOs: 2 to 5, 8 to 10, 13, and 15 to 20 from the nucleic acid sample or a polymorphic site of a complementary nucleotide thereof ( However, there is provided a method for detecting a single nucleotide polymorphism in a specimen, including the polymorphic site (as defined above).

In addition, according to another aspect of the present invention, in order to provide information necessary for diagnosing autism, (i) obtaining a nucleic acid sample from the specimen; And (ii) analyzing a haplotype from the nucleic acid sample.

Polynucleotides (b) to (e), (h) to (j), (m), (o) to (t) according to the present invention at least one polynucleotide selected from the group consisting of autism It can be usefully used for diagnosis.

In addition, a primer or probe composed of the polynucleotide or a complementary nucleotide thereof, and the polynucleotide or a complementary nucleotide kit thereof may be usefully used for diagnosis of autism.

In addition, the detection method of the present invention can effectively provide information necessary for diagnosing autism.

1 shows the linkage disequilibrium block of the MTHFR gene.
Figure 2 shows the linkage disequilibrium block of the RELN and LAMB1 genes.
3 shows a haploid type formed in comparison with the result of genotype analysis from an association unbalanced block of the MTHFR gene.
Figure 4 shows the haploid type formed in comparison with the analysis result of genotype from the linkage disequilibrium block of RELN and LAMB1 genes.

In the present specification, the term "autism" is also referred to as autism spectrum disorder (ASD), and is a pervasive developmental disorder (PDD), which is a social interaction disorder, communication disorder, and repetition. It is a generic term for neurodevelopmental disorders with homologous behavior. Autism appears in various forms and varies in severity from individual to individual.Autistic disorder, Rett's disorder, childhood disintegrative disorder, Asperger's syndrome, and general developmental disorder with specific inability ( Pervasive Developmental Disorder Not Otherwise Specified; PDD-NOS). The PDD-NOS is a disorder that is close to autism, but does not meet the diagnostic criteria of the American Psychiatric Association, Diagnostic and Statistical Manual of Mental Disorder-IV (DSM-IV). This refers to a case in which all of the symptoms do not appear and some of them appear, and is defined as atypical autism disorder in the International Classification of Disease (ICD) prepared by the WHO. In the present specification, the term "autism" also includes the PDD-NOS.

The present invention relates to autism-related polynucleotides (b) to (e), (h) to (j), (m), at least one polynucleotide selected from the group consisting of (o) to (t) or a complementary nucleotide thereof Provides. The polynucleotide or the DNA sequence of SEQ ID NOs: 2 to 5, 8 to 10, 13, and 15 to 20 from which the polynucleotide or its complementary complementary nucleotide is derived is a single nucleotide polymorphism showing a statistically significant difference between the autism patient group and the normal population. As, these are MTHFR (5,10-Methylenetetrahydrofolate reductase), AUTS2 (autism susceptibility candidate 2), RELN (reelin), LAMB1 (laminin, beta 1laminin, beta 1), WNT2 (wingless-type MMTV integration site family member 2), respectively. , FOXP2 (forkhead box P2), and ST7 (suppression of tumorigenicity 7) genes.

The MTHFR gene is a gene encoding an enzyme that converts 5,10-methylenetetrahydrofolate to 5-methyltetrahydrofolate, and the resulting methyl group is provided to homocysteine. It is involved in the metabolic process that synthesizes methionine. Mutations in this gene decrease the activity of the enzyme, causing an excess of homocysteine to accumulate in the body. This blood homocysteine hyperactivity is known to act as a cause of various vascular diseases and cancers. The AUTS2 (autism susceptibility candidate 2) gene is known as a gene susceptible to autism, but its clear function has not yet been opened. The RELN gene is a gene that encodes a secretory protease and plays an important role in the movement of nerve cells. When the concentration of the enzyme is low, it weakens the neural circuit required for normal recognition, leading to schizophrenia and autism. It is known to cause the onset. The LAMB1 gene is a gene that encodes one of the three subtypes of the beta chain among the three protein chains (laminin alpha, beta and gamma) that make up laminin, an important component of the basement membrane. The WNT2 gene is a gene involved in the release of proteins during signaling. The WNT2 gene is one of a family of 16 WNT genes that influence brain development. The FOXP2 gene is a protein that binds to DNA as one of the transcription factors, and is a gene that acts as a switch that regulates the production of other gene products. Mutations in the FOXP2 gene cause vocalization damage in other organisms, and are known to be inaccurate in pronunciation and poor comprehension of sentences. Although the function of the ST7 gene is not yet clearly known, it is known as a gene present in the region of chromosome 7 identified as an autism-susceptibility locus.

The present inventors selected MTHFR, AUTS2, RELN, LAMB1, WNT2, FOXP2, and ST7 as candidate gene groups related to autism, and conducted various studies to develop diagnostic indicators at the molecular level in diagnosing autism. MTHFR, AUTS2, RELN, LAMB1, WNT2, FOXP2, and ST7 are known to have numerous monobasic polymorphs of 223, 3158, 2755, 306, 129, 647, and 271, respectively (NCBI dbSNP; http://www. .ncbi.nlm.nih.gov/projects/SNP/). The present inventors performed genotyping and statistical analysis on 180 Korean autistic patients and 147 normal people. Surprisingly, only a small fraction of the single base polymorphisms were statistically significant differences between the autism patient group and the normal group. Was found to represent. In particular, the monobasic polymorphs of SEQ ID NOs: 2 to 5, 8 to 10, 13, and 15 to 20 have risk alleles at each polymorphic site as a result of Hardy-Weinberg Equilibrium and logistic regression analysis. Was confirmed.

Accordingly, the present invention relates to autism-related polynucleotides (b) to (e), (h) to (j), (m), one or more polynucleotides selected from the group consisting of (o) to (t) or a complement thereof Red nucleotides, and the length of the polynucleotide or its complementary nucleotide is 10 to 100 nucleotides, preferably 20 to 60 nucleotides, more preferably 40 to 60 nucleotides.

At least one polynucleotide selected from the group consisting of the polynucleotides (b) to (e), (h) to (j), (m), and (o) to (t) according to the present invention is a polymorphic sequence. A polymorphic sequence refers to a sequence including a polymorphic site representing a single nucleotide polymorphism in a nucleotide sequence. Polymorphic site refers to a site in which a single base polymorphism occurs in a polymorphic sequence. The polynucleotide can be DNA or RNA.

The present invention is also specific for autism diagnosis alleles hybridizing with 10 or more consecutive polynucleotides derived from DNA sequences of SEQ ID NOs: 2 to 5, 8 to 10, 13, and 15 to 20 including polymorphic sites or complementary nucleotides thereof. Contains red polynucleotides. The allele-specific polynucleotide means specifically hybridizing to each allele. That is, it means that it can specifically hybridize to one of the alleles represented by the monobasic polymorphism.

The present invention also provides a polynucleotide selected from the group consisting of the polynucleotides (b) to (e), (h) to (j), (m), (o) to (t), or a complementary polynucleotide thereof. Consisting of, including amplification allele-specific primers for the diagnosis of autism. Herein, "primer" refers to a template-directed DNA synthesis under appropriate conditions (eg, 4 different nucleoside triphosphates (ATP, GTP, CTP, TTP) and DNA polymerase) in an appropriate buffer. It refers to a single-stranded oligonucleotide that can serve as a starting point. The appropriate length of the primer may vary depending on the purpose of use, but is usually 10 to 100, preferably 10 to 80 nucleotides. Short primer molecules generally require a lower temperature to form a stable hybrid with the template. The primer sequence need not be completely complementary to the template, but must be sufficiently complementary to hybridize to the template. It is preferable that the 3'-end of the primer is aligned with the monobasic polymorphs of SEQ ID NOs: 2 to 5, 8 to 10, 13, and 15 to 20. The primer hybridizes to a target DNA containing a polymorphic site, and amplification is initiated from the allelic DNA in which the primer shows complete homology. This primer is used in pairs with a second primer that hybridizes to the opposite side. The product is amplified from the two primers by amplification, and only the amplified product is specifically stained and visualized. This means that certain allelic forms exist. When the allele-specific polynucleotides used herein undergo different denaturation processes, they ^{can be used not only by GoldenGate TM} Assay, but also by other methods.

The present invention consists of one or more polynucleotides selected from the group consisting of polynucleotides (b) to (e), (h) to (j), (m), (o) to (t) or complementary polynucleotides thereof, It includes a probe for autism diagnosis and a microarray for autism diagnosis including the same. In addition, the present invention is a polynucleotide selected from the group consisting of polynucleotides (b) to (e), (h) to (j), (m), (o) to (t) or a complementary polynucleotide thereof. It includes a kit for diagnosing autism, including.

The present invention provides information necessary for diagnosing autism, comprising the steps of: (i) obtaining a nucleic acid sample from a specimen; And (ii) analyzing the nucleotide sequence of a polymorphic site of a polynucleotide selected from one or more polynucleotides of SEQ ID NOs: 2 to 5, 8 to 10, 13, and 15 to 20 from the nucleic acid sample or a polymorphic site of a complementary nucleotide thereof ( However, the polymorphic site includes a method of detecting a single nucleotide polymorphism in a specimen, including).

The sample includes blood, tissue, cells, and the like, and a nucleic acid sample can be obtained according to a conventional method. For example, in the case of blood, a nucleic acid sample can be obtained as follows. The blood is transferred to a centrifugation test tube and a low-salt buffer solution (10mM Tris-HCl [pH 7.6], 10mM KCl, 10mM MgCl2, and 2mM EDTA) is added. Then, after adding Nonidet P-40, the resulting solution is well mixed, and then centrifuged at about 2,200 rpm for 10 minutes at room temperature. At this time, the obtained mass was resuspended in a high-salt buffer solution (10mM Tris-HCl [pH 7.6], 10mM KCl, 10mM MgCl2, 0.4M NaCl, and 2mM EDTA)). Genomic DNA is mixed with 50ul of 10% SDS, and then reacted at about 55°C overnight. After the reaction, each genomic DNA is transferred to a 1.5 ml centrifugal test tube, and 0.3 ml of 6M NaCl is added. After mixing the genomic DNAs well, centrifuge for 10 minutes at about 12,000 rpm. After centrifugation, the upper layer containing genomic DNA is collected and transferred to a new test tube, and an equal amount of 100% ethanol is added at room temperature. Mix well until genomic DNAs precipitate, and when precipitated, wash genomic DNAs with 70% ethanol. After this process, open the lid of the test tube and let the solution components dry. Then, a nucleic acid sample can be obtained by resuspending genomic DNA by adding a TE buffer solution (pH 8.0).

Here, the step of determining the nucleotide sequence of the polymorphic site is a polynucleotide selected from the group consisting of polynucleotides of SEQ ID NOs: 2 to 5, 8 to 10, 13, and 15 to 20 or a polymorphic site as a complementary polynucleotide sequence thereof. It may be performed using a primer or a probe using a sequence including (however, the polymorphic site is as defined above) as a template. The primer or probe is as described above.

In addition, the step of analyzing the nucleotide sequence of the polymorphic site may be a polynucleotide selected from the group consisting of polynucleotides (b) to (e), (h) to (j), (m), (o) to (t), or Hybridizing the nucleic acid sample to a microarray to which its complementary nucleotide is immobilized; And analyzing the obtained hybridization result, and the length of the DNA sequence of the polynucleotide or its complementary nucleotide immobilized on the microarray may be 10 to 100 nucleotides, respectively.

The results obtained according to the method of detecting a single nucleotide polymorphism in a specimen of the present invention can provide information necessary for autism diagnosis, for example, polymorphisms of SEQ ID NOs: 2 to 5, 8 to 10, 13, and 15 to 20. If more than one nucleotide sequence of the site is C, A, G, A, A, A, C, A, T, G, T, A, C, and G (risk alleles), respectively, in the autism patient group It can be judged as belonging. It can be determined that the more nucleic acid sequences having the risk alleles are detected in one sample, the higher the probability of belonging to the autism patient group.

In addition, analyzing the nucleotide sequence of the polymorphic site may include genotype analysis of the polymorphic sites of SEQ ID NOs: 2 to 5, 8 to 10, 13, and 15 to 20. Preferably, the step of analyzing the nucleotide sequence of the polymorphic site includes analysis of a co-dominant genotype and a recessive genotype of the polymorphic site of SEQ ID NOs: 2, 8, 10 and 16, or Including dominant genotype analysis of polymorphic sites of Nos. 3, 5 and 18, or co-dominant genotype and dominant genotype analysis of polymorphic sites of SEQ ID Nos. 4, 17 and 19 It may include, or include a recessive genotype analysis of the polymorphic site of SEQ ID NO: 9, 13 and 15, or may include a co-dominant genotype analysis of the polymorphic site of SEQ ID NO: 20.

That is, if the polymorphic sites of SEQ ID NOs: 2, 8, 10 and 16 each have one or more genotypes of C/C, A/A, C/C, and G/G, it can be determined that the risk of developing autism is high. In addition, since it is also significant in the co-dominant model, it can be judged that the risk of autism is high if the risk alleles C, A, C, and G are present. In addition, when the polymorphic sites of SEQ ID NOs: 3, 5, and 18 each have one or more genotypes of A/A, A/A, and A/A, it can be determined that the risk of developing autism is high. In addition, if the genotypes of the polymorphic sites of SEQ ID NOs: 4, 17 and 19 are G/G and A/G, T/T and C/T, and C/C and T/C, respectively, the risk of autism is increased. It can be judged to be high, and since it is also significant in the co-dominant model, it can be judged that the risk of autism is high if the risk alleles G, T, and C are present. In addition, when the polymorphic sites of SEQ ID NOs: 9, 13, and 15 each have one or more genotypes of A/A, A/A, and T/T, it can be determined that the risk of developing autism is high. In addition, when SEQ ID NO: 20 has a risk allele, G, it can be determined that the risk of developing autism increases. The more nucleic acid sequences having the risk alleles are detected in one sample, the higher the probability of belonging to the risk group can be determined.

The present invention also provides information necessary for diagnosing autism, comprising the steps of: (i) obtaining a nucleic acid sample from a specimen; And (ii) analyzing a haplotype from the nucleic acid sample. Specifically, the haplotype analysis is a haploid type determined from the polymorphic site of the DNA sequence of SEQ ID NOs: 1 and 2 from the nucleic acid sample, the haploid type determined from the polymorphic site of the DNA sequence of SEQ ID NOs: 6 and 7, or SEQ ID NO: It can be performed by analyzing the haplotype determined from the polymorphic sites of the DNA sequences of 8, 9, 11, 12, 13 and 14. The polymorphic sites of the DNA sequence of SEQ ID NOs: 1 and 2 refer to the 401th base of SEQ ID NO: 1 and the 301th base of SEQ ID NO: 2, respectively, and the polymorphic sites of the DNA sequences of SEQ ID NOs: 6 and 7 are each of SEQ ID NO: 6 It refers to the 499th base and the 301th base of SEQ ID NO: 7, and the polymorphic sites of the DNA sequences of SEQ ID NOs: 8, 9, 11, 12, 13 and 14 are the 301th base of SEQ ID NO: 8 and the 301th base of SEQ ID NO: 9, respectively. It refers to the base, the 301th base of SEQ ID NO: 11, the 301th base of SEQ ID NO: 12, the 201th base of SEQ ID NO: 13, and the 567th base of SEQ ID NO: 14.

In other words, as a result of haploid analysis determined from the polymorphic sites of the DNA sequences of SEQ ID NOs: 1 and 2, it can be determined that a person who does not have a GA haploid type or has a GC haploid type has a higher risk of autism than those who do not. . In addition, as a result of haploid analysis determined from the polymorphic sites of the DNA sequences of SEQ ID NOs: 6 and 7, it can be determined that a person who does not have a TC haploid type has a higher risk of developing autism than a person who has it. In addition, as a result of haploid analysis determined from the polymorphic sites of the DNA sequences of SEQ ID NOs: 8, 9, 11, 12, 13 and 14, it was determined that people with AAGAAG haploid type have a higher risk of autism than those who do not have it. Can (see Table 7 and Table 8, Figures 1-4).

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

Example

1. Selection of subjects

Autism patients were 7 to 22.5 years old, and 180 outpatients and students of three special schools were selected from 2004 to 2006, all of which were DSM-IV (American Psychiatric Association, 1994), clinical interview, and symptom grade. (symptom rating) was performed, and patients with a Childhood Autism Rating Scale (CARS) score of 30 or higher were selected. All diagnoses were performed by two independent experienced psychiatrists and psychologists. The control group was selected from 147 healthy subjects with no genetic or neurological etiology. After explaining the purpose and method of the study, all subjects obtained consent through written consent, and passed their Institutional Review Board (IRB). Blood was collected from a total of 327 people selected, and genomic DNA in the blood was separated and used in the experiment. The sizes of the used clusters are shown in Table 1 below.

group Normal person Patient group total Number of investigators 147 180 327

2. Preparation of genomic DNA

Blood was collected from 327 people to prepare genomic DNA (gDNA). Blood was provided with 300 ul each, and Gentra's blood genomic DNA preparation kit was used to extract genomic DNA. The prepared genomic DNA was adjusted to a concentration of 250 ng/5 ul using a ^{Quant-iT™ Picogreen dsDNA assay kit (Molecular Probes, Invitrogen) for GoldenGate Assay.}

3. Amplification and analysis of genomic DNA

According to the Goldengate assay method, genotype was analyzed using a Sentrix array matrix chip. All reagents for GoldenGate assay and Sentrix array matrix chip were purchased from Illumina Inc. and used. All procedures were performed according to the Illumina's GoldenGate assay method.

Genomic DNA samples (250 ng/5 µl) were mixed with 5 ul of GS#-MS1 reagent in a 96 well plate. The plate was sealed, centrifuged at 250×g for 1 minute, and reacted in a 95° C. heat block for 30 minutes. Here, 5 ul GS#-SUD was added, mixed, 5 ul 2-propanol (2-propanol) was added, and after mixing again, centrifugation at 3,000×g for 20 minutes, the supernatant was removed, and the pellet was dried. The dried DNA was well released with 10 ul of GS#-RS1 reagent. Add 10 ul of GS#-OPA and 30 ul of GS#-OB1, mix well, put in a 70℃ heat block, and cool slowly until it reaches 30℃. The GS#-ASE plate was placed on a magnetic plate and left for 2 minutes. The supernatant was removed, 50 ul GS#-AM1 was added and mixed well. Placed on a magnetic plate and left to stand for 2 minutes, the supernatant was removed, 50 ul GS#-AM1 was added and mixed well. After washing with 50 ul GS#-UB1 in the same manner as above, the supernatant was removed, 37 ul GS#-MEL was added, mixed well, and incubated at 45° C. for 15 minutes. The upper plate was put on the magnetic plate again, left for 2 minutes, the supernatant was removed, and 50 ul GS#-UB1 was added. The above process was repeated again, 35 ul GS#-IP1 was added, mixed well, and incubated for 1 minute in a 95°C heat block. The plate was placed on a magnetic plate and left for 2 minutes. After transferring 30 ul of the supernatant to a GS#-PCR plate, 64 ul of illumina-recommended DNA polymerase and 100 ul UDG were added to a GS#-MMP tube, mixed well, and amplified under the conditions shown in Table 2.

Temperature Time at each temperature 37℃ 10 min 95℃ 3 min 34 cycles 95℃ 35 sec 56℃ 35 sec 72℃ 2 min 72℃ 10 min 4℃ 5 min

After the reaction, 20 ul of GS#MPB was added and mixed well, transferred to a 96-well filter plate, and left in the dark for 60 minutes. The filter plate adapter was placed in a 96-well V-bottom and centrifuged to remove the supernatant. Again, 50 ul GS#-UB2 was added to the filter plate and centrifuged at 1000 x g for 5 minutes. A GS#-INT plate containing 30 ul of GS#_MH1 was placed on the filter plate, and 30 ul of 0.1N NaOH was added to the filter plate. The PCR product was collected by centrifugation at 1000 xg for 5 minutes.

The chip was pretreated with GS#-UB2 and NaOH, and the collected PCR product was transferred to 384 wells, and the pretreated chip was placed there. Hybridization was performed at 60° C. for 30 minutes, and the temperature was changed to 45° C. and hybridized for 14 hours or more. After washing in GS#UB2 and GS#IS1 and drying at room temperature, image scanning was performed for analysis.

Genotyping Analysis of the genotype from the image file was performed using Illumina's Beadstudio (version 2.3.41.16318) to confirm the genotype for each polymorphism. Beadstudio represents a typical genotypic pattern with the ratio of the intensity of color development between probes that recognize two alleles.

4. Statistical Analysis

The genotype for each single base polytype was analyzed using Haploview software (version 53.32) and SAS statistical software (SAS Institute, Cary, NC). The significance level for the test of significance was 5%, and significance was placed only when the P value was 0.05 or less.

The standard was set to have a Minor Allele Frequency (MAF) of 5% or more from each single-base polymorphism result, and the Hardy-Weinberg Equilibrium (HWE) test was performed through chi-square analysis. Single base polymorphs with a level (p value) >0.05 or higher were selected. From the results selected here, the association of disease for alleles and genotypes was obtained through Odds Ratio (OR) and tested for significant differences between the two groups. The odds ratio was calculated through a logistic regression model, and dominant, recessive, and co-dominant models were applied. In analyzing the significance of the disease according to each model, if the majority allele is A1 and the minority allele is A2, then the dominant model compares the sum of the frequencies of A1A1 and the frequencies of A1A2 and A2A2. The (Recessive) trait model compares the sum of A1A1 and A1A2 with the frequency of A2A2, and the Co-Dominant trait model compares the frequencies for A1A1, A1A2 and A2A2.

Two or more monobasic polymorphisms form a linkage disequilibrium and form a group, which is called a linkage disequilibrium block. Association unequilibrium blocks were identified by the method proposed by Gabriel (Gabriel SB, Schaffner SF, Nguyen H, Moore JM, Roy J, Blumenstiel B, et al. The structure of haplotype blocks in the human genome. Science;296:2225 -2229 (2002)). The genetic unit formed from the associated non-equilibrium blocks is determined by genotyping of each single nucleotide polymorphism, which is called a haplotype. Haploview was used to test linkage disequilibrium (LD) between single nucleotide polymorphisms showing close association at the gene level, and the association of the haplotype obtained from this to disease was tested through Oz ratio. . In addition, Rwanton's (Lewontin's) D'| and r ² measurements were used.

5. Results

(1) MTHFR, AUTS2, RELN, LAMB1, WNT2, FOXP2, and a single nucleotide polymorphism showing significance present in the ST7 gene

As a result of selecting MTHFR, AUTS2, RELN, LAMB1, WNT2, FOXP2, and ST7 as candidate gene groups related to autism, and analyzing the results obtained through the genotyping analysis, surprisingly, among many single-base polymorphisms, MTHFR, AUTS2, and WNT2 It was found that two single-base polymorphs for each gene, three single-base polymorphs for the RELN gene, nine single-base polymorphs for the LAMB1 gene, and one single polynucleotide for each of the FOXP2 and ST7 genes were statistically significant. The characteristics of the polymorphic sites of these monobasic polymorphs are shown in Table 3 below.

Sequence number Reference
SNP ID Allele Chromosomal region location gene SNP role Amino acid changes One rs2274976 G>A 1p36.3 11785193 MTHFR exon11 Gln->Arg 2 rs1801131 A>C 1p36.3 11788742 MTHFR exon7 Ala->Glu 3 rs736477 A>G 7q11.22 69570924 AUTS2 intron No change 4 rs2293503 A>G 7q11.22 69694776 AUTS2 intron No change 5 rs13232021 A>G 7q22 102735098 RELN intron No change 6 rs362646 T>C 7q22 102869754 RELN intron No change 7 rs2072402 C>A 7q22 102885872 RELN intron No change 8 rs7561 C>A 7q22 107158317 LAMB1 3'UTR No change 9 rs7797759 C>A 7q22 107158958 LAMB1 intron No change 10 rs13646 T>C 7q22 107159089 LAMB1 intron No change 11 rs2237685 G>A 7q22 107168735 LAMB1 intron No change 12 rs10271464 G>A 7q22 107173554 LAMB1 intron No change 13 rs2158836 G>A 7q22 107174790 LAMB1 intron No change 14 rs740287 T>G 7q22 107178731 LAMB1 intron No change 15 rs4730283 C>T 7q22 107187940 LAMB1 exon22 Arg->Gln 16 rs1548638 G>T 7q23 107198769 LAMB1 intron No change 17 rs936145 C>T 7q31 113891131 FOXP2 intron No change 18 rs12706126 A>G 7q31.1-q31.3 116185759 ST7 5'-flanking
UTR No change 19 rs6948009 T>C 7q31 116510364 WNT2 3'-flanking
UTR No change 20 rs3779548 A>G 7q31 116524846 WNT2 intron No change

ㆍThe sequence number is a sequence number representing each single nucleotide polymorphism.

ㆍReference SNP ID means the name of the standard single nucleotide polymorphism (Reference SNP ID; rs), which is to distinguish each single nucleotide polymorphism in the database of the National Institute of Health Biotechnology Information Research (NCBI) after the Human Genome Project. This is the name I gave.

ㆍThe chromosome area represents the unit divided according to color when the chromosome is stained with a specific technique. This is called the G-band.

ㆍLocation means the position of the standard base sequence on the chromosome (NCBI Genome build 36.2).

• Gene indicates the gene in which the single nucleotide polymorphisms are located.

The role of SNP indicates where the single nucleotide polymorphisms are located in the gene. Thus, it is possible to see how each single polymorphism works in gene expression and function.

ㆍAmino acid change is an explanation of whether the above monobasic polymorphisms change the amino acids constituting the protein when the gene is expressed to produce a protein.

(2) Genotype and risk allele analysis

Table 4 shows the frequency of minor alleles of SEQ ID NOs: 1 to 20 and the number and frequency of each genotype, and Tables 5 and 6 show that for each genotype frequency of Table 4, each single nucleotide polymorphism is a dominant model, a recessive ( Recessive) model and co-dominant (incomplete dominant) model, and the results of testing whether there is a relationship with autism and risk alleles are shown. Risk alleles are alleles with a high risk of developing autism if they have these alleles.

Sequence number Opposition
factor Minority allele frequency A1A1 A1A2 A2A2 Control Autism Control Autism Control Autism Control Autism One G>A 0.09 0.10 123(83.67) 145 (80.56) 23(15.65) 35(19.45) 1(0.68) - 2 A>C 0.20 0.28 94(63.95) 96(54.24) 48(32.65) 64(36.16) 5(3.40) 17(9.60) 3 A>G 0.09 0.05 122(82.99) 163 (90.56) 24(16.33) 16(8.89) 1(0.68) 1(0.56) 4 A>G 0.38 0.49 54(36.73) 42(23.33) 73(49.66) 99 (55.00) 20(13.61) 39(21.67) 5 A>G 0.27 0.23 76(51.70) 113(62.78) 63(42.86) 52(28.89) 8(5.44) 15(8.33) 6 T>C 0.42 0.49 48(32.65) 43(23.89) 75(51.02) 97(53.89) 24(16.33) 40(22.22) 7 C>A 0.14 0.14 109 (74.15) 133 (73.89) 34(23.13) 45(25.00) 4(2.72) 2(1.11) 8 C>A 0.13 0.19 110(74.83) 122 (67.78) 36(24.49) 48(26.67) 1(0.68) 10(5.56) 9 C>A 0.13 0.19 97(74.05) 121 (68.75) 33(25.19) 44(25.00) 1(0.76) 11(6.25) 10 T>C 0.13 0.20 110(74.83) 123(68.33) 36(24.49) 42(23.33) 1(0.68) 15(8.33) 11 G>A 0.30 0.30 73(49.66) 88(48.89) 61(41.50) 76(42.22) 13(8.84) 16(8.89) 12 G>A 0.14 0.18 108(73.47) 127 (70.56) 37(25.17) 43(23.89) 2(1.36) 10(5.56) 13 G>A 0.13 0.18 109 (74.15) 126 (70.00) 37(25.17) 44(24.44) 1(0.68) 10(5.56) 14 T>G 0.15 0.20 105 (71.43) 119(66.11) 39(26.53) 51(28.33) 3(2.04) 10(5.56) 15 C>T 0.14 0.20 107 (72.79) 117(65.36) 38 (25.85) 51(28.49) 2(1.36) 11(6.15) 16 G>T 0.32 0.24 71(48.30) 101(56.11) 59(40.14) 70(38.89) 17(11.56) 9(5.00) 17 C>T 0.07 0.12 127(86.39) 137 (76.11) 18(12.24) 42(23.33) 2(1.36) 1(0.56) 18 A>G 0.20 0.14 93(63.27) 134 (74.44) 50(34.01) 41(22.78) 4(2.72) 5(2.78) 19 T>C 0.15 0.23 106 (72.11) 105(58.33) 39(26.53) 68 (37.78) 2(1.36) 7(3.89) 20 A>G 0.34 0.41 60(40.82) 56(31.11) 74(50.34) 100 (55.56) 13(8.84) 24(13.33)

HWE and logistic regression analysis of each single base polymorphism in autism patients and normal people Sequence number HWE model OR (95% CI) p-value Risk antagonist One Control 0.09 Kong Woosung - - - Autism 0.15 dominant - - zeal - - 2 Control 0.71 Kong Woosung 1.53 (1.06∼2.20) 0.023 C Autism 0.20 dominant 1.50 (0.96∼2.34) 0.078 zeal 3.02 (1.09-8.39) 0.034 3 Control 0.88 Kong Woosung 0.55 (0.30∼1.02) 0.058 A Autism 0.39 dominant 0.51 (0.26∼0.98) 0.045 zeal 0.81 (0.05∼13.13) 0.885 4 Control 0.55 Kong Woosung 1.61 (1.16∼2.24) 0.005 G Autism 0.18 dominant 1.91 (1.18∼3.09) 0.009 zeal 1.76 (0.97-3.17) 0.061 5 Control 0.27 Kong Woosung 0.81 (0.57∼1.15) 0.240 A Autism 0.02 dominant 0.63 (0.41∼0.99) 0.044 zeal 1.58 (0.65∼3.84) 0.313 6 Control 0.56 Kong Woosung 1.37 (0.99∼1.90) 0.055 - Autism 0.29 dominant 1.54 (0.95∼2.51) 0.079 zeal 1.46 (0.84∼2.57) 0.183 7 Control 0.50 Kong Woosung 0.94 (0.60∼1.48) 0.803 - Autism 0.40 dominant 1.01 (0.62∼1.67) 0.957 zeal 0.40 (0.07∼2.22) 0.296 8 Control 0.29 Kong Woosung 1.54 (1.01∼2.35) 0.047 A Autism 0.08 dominant 1.41 (0.87∼2.30) 0.163 zeal 8.59 (1.09∼67.89) 0.041 9 Control 0.31 Kong Woosung 1.45 (0.94∼2.24) 0.090 A Autism 0.02 dominant 1.30 (0.78∼2.15) 0.312 zeal 8.67 (1.10∼67.99) 0.040 10 Control 0.29 Kong Woosung 1.58 (1.06∼2.37) 0.026 C Autism 0.00 dominant 1.38 (0.85∼2.24) 0.197 zeal 13.27 (1.73∼101.64) 0.013

HWE and logistic regression analysis of each single base polymorphism in autism patients and normal people Sequence number HWE model OR (95% CI) p-value Risk antagonist 11 Control 0.96 Kong Woosung 1.02 (0.73∼1.43) 0.910 - Autism 0.94 dominant 1.03 (0.67-1.59) 0.890 zeal 1.01 (0.47∼2.16) 0.989 12 Control 0.56 Kong Woosung 1.28 (0.85-1.94) 0.237 - Autism 0.02 dominant 1.16 (0.71∼1.88) 0.560 zeal 4.26 (0.92-19.78) 0.064 13 Control 0.26 Kong Woosung 1.39 (0.91∼2.11) 0.129 A Autism 0.03 dominant 1.23 (0.75∼2.00) 0.407 zeal 8.59 (1.09∼67.89) 0.041 14 Control 0.78 Kong Woosung 1.34 (0.90∼2.00) 0.154 - Autism 0.16 dominant 1.28 (0.80∼2.06) 0.304 zeal 2.82 (0.76∼10.46) 0.120 15 Control 0.50 Kong Woosung 1.50 (1.00∼2.26) 0.050 T Autism 0.10 dominant 1.42 (0.88∼2.28) 0.151 zeal 4.75 (1.04∼21.76) 0.045 16 Control 0.38 Kong Woosung 0.70 (0.50-0.99) 0.044 G Autism 0.48 dominant 0.73 (0.47∼1.13) 0.160 zeal 0.40 (0.17∼0.93) 0.034 17 Control 0.16 Kong Woosung 1.74 (1.01∼2.99) 0.047 T Autism 0.24 dominant 1.99 (1.11∼3.57) 0.020 zeal 0.41 (0.04∼4.51) 0.462 18 Control 0.37 Kong Woosung 0.67 (0.44∼1.02) 0.059 A Autism 0.40 dominant 0.59 (0.37∼0.95) 0.030 zeal 1.02 (0.27-3.87) 0.975 19 Control 0.45 Kong Woosung 1.79 (1.17∼2.73) 0.007 C Autism 0.32 dominant 1.85 (1.16∼2.94) 0.010 zeal 2.93 (0.60-14.34) 0.184 20 Control 0.14 Kong Woosung 1.42 (1.00∼2.01) 0.047 G Autism 0.05 dominant 1.53 (0.97∼2.41) 0.069 zeal 1.59 (0.78∼3.24) 0.205

ㆍHWE state represents the state of Hardy-Weinberg Equilibrium. In the chi-square (df=1) test, based on chi-value = 3.84 (p-value=0.05, df=1), if it is greater than 3.84, it is judged as Hardy-Weinberg Equilibrium (HWE), If it was less than 3.84, it was judged as Hardy-Weinberg Disequilibrium.

• Odds ratio (OR) represents the ratio of odds to have risk alleles in the normal group to odds to have risk alleles in the patient group. The 95% CI represents the 95% confidence interval of the odds ratio and represents (lower confidence interval, upper confidence interval). If the confidence interval contains 1, the association with disease cannot be judged to be significant. If the odds ratio is greater than 1 and the 95% confidence interval does not contain 1, minority alleles have a higher frequency in the disease group, and when the odds ratio is less than 1, majority alleles have a higher frequency in the disease group. Have.

ㆍRisk alleles were considered as minority alleles when the odds ratio was greater than 1, and majority alleles were used when the odds ratio was less than 1.

Allele can determine the degree of association (risk) of the risk allele with the disease through the ratio of the frequency of occurrence of alleles between the patient group and the normal group. Since there is no significance when 1 is included in the 95% confidence interval, as can be seen from Tables 5 and 6, SEQ ID NOs: 1, 6, 7, 11, 12, and 14 do not have significance. . Accordingly, the nucleotide sequences of the polymorphic sites of SEQ ID NOs: 2 to 5, 8 to 10, 13, and 15 to 20 are respectively C, A, G, A, A, A, C, A, T, G, T, A, If more than one of C and G (risk alleles) is present, it can be determined that the possibility of developing autism is high.

In addition, SEQ ID NOs: 2, 8, 10, and 16 show significance in the co-dominant and recessive models, and SEQ ID NOs: 2, 8, and 10 have an odds ratio> 1, and the genotype has a minor allele. People with a major allele have a higher risk of developing autism than those with the Major Allele. SEQ ID NO: 16 shows that people with an odds ratio <1 and a genotype of multiple alleles have a higher risk of autism than those with a small number of alleles.

SEQ ID NOs: 3, 5, and 18 showed significance in the dominant model, and it was found that people with an odds ratio of <1 and a genotype of multiple alleles have a higher risk of autism than those consisting of two minor alleles.

SEQ ID NOs: 4, 17 and 19 showed significance in the co-dominant and dominant model, and it was found that people with an odds ratio >1 and genotypes with minority alleles have a higher risk of autism than those with two majority alleles.

SEQ ID NOs: 9, 13, and 15 show significance in the recessive model, and an odds ratio of >1 indicates that people with two minor alleles have a higher risk of autism than those with one or more multiple alleles. . SEQ ID NO: 20 shows significance in the co-dominant model, and the odds ratio is >1, indicating a higher risk of autism with minority alleles.

From the above results, it is judged that the polymorphic sites of SEQ ID NOs: 2, 8, 10 and 16 have a high risk of autism when one or more genotypes are C/C, A/A, C/C, and G/G respectively. It can be done, and it is also significant in the co-dominant model, so if you have risk alleles C, A, C, and G, you can judge that your risk of developing autism is high. In addition, when the polymorphic sites of SEQ ID NOs: 3, 5, and 18 each have one or more genotypes of A/A, A/A, and A/A, it can be determined that the risk of developing autism is high. In addition, if the genotypes of the polymorphic sites of SEQ ID NOs: 4, 17 and 19 are G/G and A/G, T/T and C/T, and C/C and T/C, respectively, the risk of autism is increased. It can be judged to be high, and since it is also significant in the co-dominant model, it can be judged that the risk of autism is high if the risk alleles G, T, and C are present. In addition, when the polymorphic sites of SEQ ID NOs: 9, 13, and 15 each have one or more genotypes of A/A, A/A, and T/T, it can be determined that the risk of developing autism is high. In addition, when SEQ ID NO: 20 has a risk allele, G, it can be determined that the risk of developing autism increases. The more nucleic acid sequences having the risk alleles are detected in one sample, the higher the probability of belonging to the risk group can be determined.

SEQ ID NO: 1 was not analyzed because a genotype consisting of two minor alleles was not found in the autism patient group, and SEQ ID NOs: 6, 7, 11, 12, and 14 were found to be unrelated, but the above single containing SEQ ID NO: 3 The base polymorphisms obtained a significant haplotype as a result of analyzing the association by confirming the haploid type from the linkage disequilibrium (see FIGS. 1 to 4).

(3) Haploid type analysis

Tables 7 and 8 and FIGS. 1 and 2 show the association between the polymorphic sites for SEQ ID NOs: 1 to 20, and FIGS. 3 and 4 are based on this, and the Linkage Disequilibrium Block formed from FIGS. 1 and 2 It represents the haplotype formed in comparison with the analysis result of the genotype from. Table 9 shows the results of the association test with autism for the haploid type of each block.

｜D'｜
block block1 block order
number One 2 r ² block1 One 0.973 2 0.294

｜D'｜
block block2 block3 block order
number 6 7 8 9 11 12 13 14 r ² block2 6 One - - - - - - 7 0.137 - - - - - - block3 8 - - 0.976 One 0.965 0.988 0.965 9 - - 0.941 0.936 0.951 0.975 0.963 11 - - 0.082 0.075 One One 0.88 12 - - 0.911 0.883 0.08 0.988 One 13 - - 0.944 0.918 0.079 0.966 One 14 - - 0.835 0.827 0.071 0.877 0.867

Associative disequilibrium block Haplotype
Control (%) Autism (%) Model OR (95% CI) p-value Block1 ht1 ht/ht 94(63.95) 95 (53.67) Kong Woosung 1.55 (1.08∼2.24) 0.018 G-A -/ht 48(32.65) 65 (36.72) dominant 1.53(0.98∼2.40) 0.062 -/- 5(3.40) 17 (9.60) zeal 3.02 (1.09-8.39) 0.034 ht2 ht/ht 1(0.68) 6 (3.39) Kong Woosung 0.54 (0.34∼0.86) 0.009 G-C -/ht 31(21.09) 54 (30.51) dominant 0.20(0.02∼1.64) 0.133 -/- 115 (78.23) 117 (66.10) zeal 0.54 (0.33-0.89) 0.017 ht3 ht/ht 1(0.68) - Kong Woosung 0.93(0.53∼1.64) 0.805 A-C -/ht 23(15.65) 32 (18.08) dominant - - -/- 123(83.67) 145 (81.92) zeal 0.88 (0.49∼1.58) 0.678 Block2 ht4 ht/ht 24 (16.33) 40 (22.22) Kong Woosung 0.73(0.53∼1.01) 0.055 C-C -/ht 75 (51.02) 97 (53.89) dominant 0.68(0.39∼1.20) 0.183 -/- 48 (32.65) 43 (23.89) zeal 0.65(0.40∼1.05) 0.079 ht5 ht/ht 30 (20.41) 21 (11.67) Kong Woosung 1.33 (0.96∼1.83) 0.082 T-C -/ht 69 (46.94) 92 (51.11) dominant 1.94 (1.06∼3.56) 0.032 -/- 48 (32.65) 67 (37.22) zeal 1.22 (0.77∼1.93) 0.390 ht6 ht/ht 4 (2.72) 2 (1.11) Kong Woosung 1.06 (0.68∼1.65) 0.803 T-A -/ht 34 (23.13) 45 (25.00) dominant 2.49 (0.45∼13.79) 0.296 -/- 109 (74.15) 133 (73.89) zeal 0.99 (0.60∼1.62) 0.957 Block3 ht7 ht/ht 45 (30.61) 47 (26.11) Kong Woosung 1.27 (0.92∼1.74) 0.142 C-C-G-G-G-T -/ht 77 (52.38) 90 (50.00) dominant 1.25 (0.77∼2.02) 0.368 -/- 25 (17.01) 43 (23.89) zeal 1.53 (0.88-2.65) 0.129 ht8 ht/ht 12 (8.16) 16 (8.89) Kong Woosung 1.01(0.72∼1.41) 0.975 C-C-A-G-G-T -/ht 58 (39.46) 68 (37.78) dominant 0.91(0.42∼1.99) 0.816 -/- 77 (52.38) 96 (53.33) zeal 1.04 (0.67∼1.61) 0.864 ht9 ht/ht 1 (0.68) 10 (5.56) Kong Woosung 0.74 (0.49∼1.14) 0.172 A-A-G-A-A-G -/ht 36 (24.49) 41 (22.78) dominant 0.12(0.01∼0.92) 0.041 -/- 110 (74.83) 129 (71.67) zeal 0.85 (0.52-1.39) 0.521

As can be seen in FIGS. 1 and 2, SEQ ID NOs: 1 and 2 form an association non-equilibrium block (block1), and SEQ ID NOs. 6 and 7 form an association non-equilibrium block (block2). In addition, linkage disequilibrium blocks (block3) were formed in SEQ ID NOs: 8, 9, 11, 12, 13 and 14. 3 and 4, the block (block1) consisting of SEQ ID NOs: 1 and 2, which is the first block, has a haploid type in the form of GA (ht1), GC (ht2) and AC (ht3), and the second block, SEQ ID NO. Block 2 consisting of 6 and 7 was represented by a haploid type of CC (ht4), TC (ht5) and TA (ht6) form, and a third block consisting of SEQ ID NOs: 8, 9, 11, 12, 13 and 14 In (block3), a haploid type was shown in the form of CCGGGT(ht7), CCAGGT(ht8), and AAGAAG(ht9). 3 and 4, there is a high frequency in which the upper haplotype appears in each block. It indicates the value and r ² | Tables 7 and 8 is shown that by evaluating the dense for each single nucleotide polymorphism associated with non-equilibrium between the blocks shown in Figs. 1 and 2 | D '. Table 9 shows the results of testing whether there is a relationship with autism according to the dominant model, the recessive model, and the co-dominant (incomplete dominant) model. In block 1, the ht1(GA) and ht2(GC) haplotypes showed significance in the co-dominant and recessive models, respectively, showing that people who do not have haploid types ht1 and ht2 have a higher risk of autism than those who have them. The ht5 (TC) in block 2 and the ht9 (AAGAAG) haplotype in block 3 showed significance in the dominant model, showing that people with ht5 and ht9 as haploid types, respectively, had a higher risk of autism than those who did not.

From the above results, as a result of haploid analysis determined from the polymorphic sites of the DNA sequences of SEQ ID NOs: 1 and 2, it is determined that the case of not having the GA haploid type or the case of having the GC haploid type is at a higher risk of developing autism than those who do not. I can. In addition, as a result of haploid analysis determined from the polymorphic sites of the DNA sequences of SEQ ID NOs: 6 and 7, it can be determined that a person who does not have a TC haploid type has a higher risk of developing autism than a person who has it. In addition, as a result of haploid analysis determined from the polymorphic sites of the DNA sequences of SEQ ID NOs: 8, 9, 11, 12, 13 and 14, it was determined that people with AAGAAG haploid type have a higher risk of autism than those who do not have it. I can.

<110> College of Medicine Pochon CHA University Industry-Academic Cooperation Foundation <120> Polynucleotides comprising single nucleotide polymorphism, microarrays and diagnostic kits comprising the same, and detection methods using the same <160> 20 <170> KopatentIn 1.71 <210> 1 <211> 801 <212> DNA <213> Homo sapiens <400> 1 cttccctggg cgagagatca tccagcccac cgtagtggat cccgtcagct tcatgttctg 60 gaaggtaaag gagccggggg caagcttgcc ccgcccacct ggaaaaccgt ggggagggat 120 tgggaccaag tcccaagcgt gtgctgaagg ccacactgga cccagccttc agggcacacc 180 cagctctgac tcacccatgt cactgctgat gcagggtgtt tatttctggg caggggtggg 240 aagtgatact ggcagtgggc cttgttctat tccgggaaat gtcctgttga gcagagccct 300 tggagagccc tgttaatctt gcctctgtgt gtgtgtgcat gtgtgcgtgt gtgcgggggt 360 atgtgtgtgt aggacgaggc ctttgccctg tggattgagc rgtggggaaa gctgtatgag 420 gaggagtccc cgtcccgcac catcatccag tacatccacg acaactactt cctggtcaac 480 ctggtggaca atgacttccc actggacaac tgcctctggc aggtggtgga agacacattg 540 gagcttctca acaggcccac ccagaatgcg agagaaacgg aggctccatg accctgcgtc 600 ctgacgccct gcgttggagc cactcctgtc ccgccttcct cctccacagt gctgcttctc 660 ttgggaactc cactctcctt cgtgtctctc ccaccccggc ctccactccc ccacctgaca 720 atggcagcta gactggagtg aggcttccag gctcttcctg gacctgagtc ggccccacat 780 gggaacctag tactctctgc t 801 <210> 2 <211> 601 <212> DNA <213> Homo sapiens <400> 2 tgcagacctt ccttgcaaat atatctttgt tcttgggagc gggagggcag aagaagtttg 60 catgcttgtg gttgacctgg gaggagtcag gggcagaatt tacaggaatg gcctcctggg 120 catgtggtgg cactgccctc tgtcaggagt gtgccctgac ctctgggcac ccctctgcca 180 ggggcaattc ctcttcccct gcctttgggg agctgaagga ctactacctc ttctacctga 240 agagcaagtc ccccaaggag gagctgctga agatgtgggg ggaggagctg accagtgaag 300 maagtgtctt tgaagtcttt gttctttacc tctcgggaga accaaaccgg aatggtcaca 360 aagtgagtga tgctggagtg gggaccctgg ttcatcccct gcccctggcc tgaccccagc 420 tgcaggccag gctgcggggc tgtgacttcc ccatcctgtg ccctcccctc catgctgtgg 480 acatggcaaa gggagaaggg taagttggga gacctccacc tggaagggct tagggaggca 540 aagacaggct gggtctttgt tgggggccgt gagagggact cagggtgcca aacctgatgg 600 t 601 <210> 3 <211> 601 <212> DNA <213> Homo sapiens <400> 3 agacactatt attcccattt cacagataaa gctaaggaag ttgcccaagg tcacagactg 60 atcaggtagc ggagctacaa cagcaagatc tgagtccaaa gcgtgtgtgt ttcaggatgt 120 cctcacactg ccaccctcct cccaccctgg tcacctgcca cagggagcga ctattcactc 180 aacagtcctc ctgaattttc ctgcctgcat ctcaaccctc attttcacat tttccagggg 240 actaccttcc cctcattcct gcaacctaat tctacctgtg aagcccccta cgctttgaag 300 ygcatccaag aaacaccttc ctgaagaaga tggcctcaga cgccgccagg gagagtttgt 360 gtttcttctg aaacaaccta tgttctgcct ggaatttcag ctgcttctgt cctggtcttc 420 tttctcctgt caggtaatta ggcggcatct agaatgccag ctaatctcca ttcaaatgcc 480 ggctctggag tgaatgataa cagtccactg tctgtgctga tgttctagta ggggagaaag 540 acaaataaaa gtcaacaaat gaataaagaa ggtaattgca gggtgtgaca tgtgtcatga 600 g 601 <210> 4 <211> 801 <212> DNA <213> Homo sapiens <400> 4 aatgcctgaa tctgcctgta cacttgcaga tgggatcgaa tacacaaagg ccacacaaag 60 catgcacctt tattaggaag agactcattt atcggggcag tgagatgtag aataactcct 120 ctccataacg tcagcttgtg tttctttccg cttcctagag gtgacctttg ctatggggca 180 agagctcacc atcttcttcc aatgggcagg aacaggctgt tgtaaactcg gattccctga 240 gataccaagg aatggctctc acccactgca ccatgggaag ttcaggcaga accactgccg 300 agcacagctc tactaccgaa agaatgcggt gtaagagatg aaatctggaa gctggacggg 360 atgtactgaa actcctaaga gagcaccatc tcctgcctaa rtggtaagca gggtatctgg 420 gactcccacc agcttgcccg gaggtgcagc gggagcatgg taaacacagc agggttgcct 480 gaagcacaca tctcagttcc cattattttg ggcgactcac ctaggtgggc agcagggttg 540 aggaagttgc tgtgatgagg aggtggccca aaaggggtcc cagttgggtg tgcagcacct 600 gttaagtcaa acataacgaa accagagttt acaaaaagcc aacttcgaag atgcaaccgg 660 cttctacttg cttaagggcc ccgtgagaaa atatgttcaa gggaggtcat ggctgaagtc 720 cacccaggcc cctgcaggca agacacaggg taaaagaaaa tgtgtgtttt tttttttttt 780 tttttttttt tttttttagg a 801 <210> 5 <211> 401 <212> DNA <213> Homo sapiens <400> 5 agtatttatc atttgtaatc catatagtct ggctcatttt atgaactgga taaactgtac 60 tagaacttga tcctaatatt cttgtactta ggaaacataa gacattttta aagtttctat 120 tcaaaggatt ttctttcctg ggggacccaa aggacattgt tatagattat gtctcataca 180 taagttggtt cctaggcatg rcccatgatg tgagtaatgc gtcttgtcca gggctttacc 240 cttaccttca gagtgtgggt caagtagcag tttggccctg agtatcccgg atcacagatg 300 cactgttccc ttaagcaatc tccatggccc ctgcagttgt ccaagcatcc agggcccagg 360 tagaaattat cgattgccca ctgcatgtct gagtacttct g 401 <210> 6 <211> 699 <212> DNA <213> Homo sapiens <400> 6 tacttttctc tctttttttt ttgagacagt ctcgctctgt cgccaggttg gagtacaatg 60 gtgcaatctc ggctcactgc aatctctgcc tgccgggttc aagcaattct cctgcctcag 120 cctcctgagt agctgggact acaggtgcgc accaccacac ccagcttttt ttttcgtatt 180 tttagtagag acagggtttc accatgttga ccaggatggt ctcaatctcc tgacctcgtg 240 atctgcctgc ctcggcctcc caaactgctg ggattacagg catgagccac tgcgcccagc 300 ctacttttct cttgatgctt ataagcatca aaatatttcc cttagatcca tatttaaaac 360 tatgacaata aattccttca aatgatcaga agtcagtggg acaaatggat gggttggctt 420 ggtagttttt cactgcaaga cacatcaatc tttgtgcgct ccatctgtca atggaaaaat 480 atctagaatt ttataccayt tgaattatga tttccccagg agaagatttc actctacaac 540 catttttcca aatgctttcc aacatacctg ggctcatgat agctgagata tgaataatgc 600 tccaggagtt tccaagttat cccattatca taagaataat gcaacaaaac tccttcacca 660 ggctgatcag gggctctgca cgtgctcaga acagatttg 699 <210> 7 <211> 601 <212> DNA <213> Homo sapiens <400> 7 cttattctgt aatgtggcaa gtattttata taatcatctt taattctcct aatagccccc 60 aatgaggtat attaaatctt taattataca gaaaaaagtg gaaagtcagg gagagtaaat 120 atgattacac agaatggaat ccaggtccaa tttcaaaatt taactttctt ccattttttc 180 tatgctgcat ttctctataa gtagcgtggc aataaccaca gaactgccat atagaattta 240 agttcatgca gtccaaaata taagtcaagt acataaagaa tgaatggcct tacaaatgtt 300 mattgtaatc ttgcatccag tttaccaaag atactatctt tctttctttc ttttttacaa 360 gaccactttt aaagaggtta ggtttcttac ctattatgac aaaaatgcta atttcaggat 420 aactcattga tttttattct gcaacatgga ttcaagtatt tcaatatctc agagagaatg 480 tactcaccca ctgtagtttt cagaggagta gacagtgctg tgggggaggt ggggtccagc 540 acagatctca ggtaagcatt cagtgtgaag gagggaccag gagcgcccat ggttggtaga 600 a 601 <210> 8 <211> 601 <212> DNA <213> Homo sapiens <400> 8 tttcacacag gcttattttt taaattactt taaggcatcc atagtttatt taaaagtgaa 60 aaatatattc actttgtctt gagatcatca tagtataaat tgctcagact ttatatatat 120 atatatctcc acagatttag aaagaaaata tgaagacaat caaagatact tagaagataa 180 agctcaagaa ttagcaagac tggaaggaga agtccgttca ctcctaaagg atataagcca 240 gaaagttgct gtgtatagca catgcttgta acagaggaga ataaaaaatg gctgaggtga 300 mcaaggtaaa acaactacat tttaaaaact gacttaatgc tcttcaaaat aaaacatcac 360 ctatttaatg tttttaatca cattttgtat ggagttaaat aaagtacagt gcttttgtat 420 atattttggt gtacttgtta ctttcactga ttgttgatga atcccttttt tcactgtaaa 480 cgtggattta taaacaagtt tactgcgtcc atgaagaccc taatttatgt tgtctataaa 540 caagctatta tatttatttg acaggtaaac atcctccaaa actccacacc cttgactccc 600 c 601 <210> 9 <211> 601 <212> DNA <213> Homo sapiens <400> 9 gctaaaagag tttttgcttc attttgtagc atttcggctt tccttctggc atcagctgac 60 tcttcagttt ttttggcaat taaattttct acttttttat acttttcatc aagttcacca 120 tctaaagtct atagttccac atttagacag aaagaagtgg taatgttagt gtcagtaatt 180 acatttaaga gcaatagtgt aatcatggca tgcattctgg attaagggcc accaaggata 240 ttcactggta agtaatttaa aattttatct tcaaacagtg gaattgaatg caaaattgac 300 mtgattttaa agaacatttc aaaaaattct ggagttgcag caaatcctta gggctaagtc 360 ctcattctcc tgattattac ttttctatag aaaagttgta gcataattgt attgcaaatt 420 acatagagaa gcgcagattt ttcaaaagta gattagacat tagagaagcc catattgaag 480 aagtggaggc tctttggatc ttgactaaat taggatttgt gaagacttgc ttataaggaa 540 atcagaggaa aaagattaaa caaaagatac aggaaggaaa ggtgagattg tagcatatgc 600 a 601 <210> 10 <211> 601 <212> DNA <213> Homo sapiens <400> 10 tagttccaca tttagacaga aagaagtggt aatgttagtg tcagtaatta catttaagag 60 caatagtgta atcatggcat gcattctgga ttaagggcca ccaaggatat tcactggtaa 120 gtaatttaaa attttatctt caaacagtgg aattgaatgc aaaattgacc tgattttaaa 180 gaacatttca aaaaattctg gagttgcagc aaatccttag ggctaagtcc tcattctcct 240 gattattact tttctataga aaagttgtag cataattgta ttgcaaatta catagagaag 300 ygcagatttt tcaaaagtag attagacatt agagaagccc atattgaaga agtggaggct 360 ctttggatct tgactaaatt aggatttgtg aagacttgct tataaggaaa tcagaggaaa 420 aagattaaac aaaagataca ggaaggaaag gtgagattgt agcatatgca agaagtgagt 480 gaggagaaga gaaatggggt gtggggttgg aatgcaaatg attcttttta gaaactgttc 540 aaatagccat ttatggaaat ttcatttcaa gagattttgt tcctttctct tgtgataaaa 600 t 601 <210> 11 <211> 601 <212> DNA <213> Homo sapiens <400> 11 gatggagaga attggcattt catgcttcat aactttgtct cctgttagct gcaatttaca 60 cattgctcta agccttcatt tttctttttt cttcatcttt tatttttcca tttaataaat 120 gacagattat ttgctcccta tttgctataa agtttcagaa gataatgagt atgaaaccta 180 gcaggcatca ataaacactg tcttatattg tgaccaattt tgtctatttc agagaaagtg 240 atgagaggag ggaggaagca tcactaagac aaagctgtaa tgaagtttga actcttctgc 300 rcctatgtac cttccatctt taactttctc tgtgtctctg ggctaagaga acacaaacct 360 tagtctataa aaaataccct ccaaattatt tcacataggt atccttcctt gaaaacacag 420 gaaggaagag gaagaaacag gctcagttag gcttttttgg ggccttctgc ctgattttca 480 ttaaactcac aacccaattt catttggctc cagagttcct ttttaaaagc ttctgggaaa 540 tgaatctact ttctaggctg gtttcagggt taatcttcac caggtaaagt tactggaaca 600 a 601 <210> 12 <211> 601 <212> DNA <213> Homo sapiens <400> 12 tgacatcaag gtccagttac tttaaatcta ggtctaggaa ggacctcctt caagtcgctt 60 gagctaatgt cagctgtcct ctgctactca tgcctggtat gagatgaaaa gtccctctgg 120 cttcttaaag gtgctgatta aagtctcatc acacacaaaa aatgttactg tcatttcgtc 180 atataaacac ctcagtgtgc aggatttaac tgtgaagcag tcccctcagg cctagatggt 240 gagtctgcag tgaaaaaacc atagcatcta gggaagcaca gatgcctcat acccttttct 300 rtttttgtcc tttagcactt aggtcttatt gtgctgtttt gggggttcct ttttggcaaa 360 aggagttggg atgtttaggg gcaagagagg ggaggtagga gaaagggtag ataaatgatc 420 cttgagaagg tcaattctct aggcttccac taacaaacac tgtccaatca gagggaagat 480 tacctagaga aggggtggga tgaggacttt tcagcataga gcagctgtta taactagatg 540 aatgaagcac tggtctggtg gactaatggc ggagaagcag aaaaaacttg aagtaagcag 600 t 601 <210> 13 <211> 617 <212> DNA <213> Homo sapiens <400> 13 agagagcaaa gcactggtgg cagggtgtgc agtcagggaa gacccccgag taccctcgcg 60 tgcacttgtc acagcgtgga ccctcaacac cctcaacgca gacacactgg cccgtggact 120 ggtcacactg tggcgtctca atgcccctgg ggtcacagtc acaggctaga agggaataag 180 caatgctagc tgatctactt ratcatgaac ctcagtgaca agatgtggca tttagaatac 240 tcacttcact gtgaaaatgg gaaggtgtca accatcgaca acaattgacc aatggctcag 300 gtagagactg tagcagcttc ctctagcaca tgcgtgactt cagtggttgc taaccccttc 360 ttagcagtaa taatacttta aacagaaatg ttttatatga atgatcccat ttaactctca 420 caataaccct atgggatacg tgttataata atcactattt cacagatgtg ggaactcaga 480 gatcagagag attatgtaac atgccccaag tctcactgca aggaaaaggc cacgccagaa 540 ttctgattca gttctttgca actccggagt ctatactctt cacctccgtc ctgcctttgc 600 ctccccctta cttagga 617 <210> 14 <211> 809 <212> DNA <213> Homo sapiens <400> 14 tgtttctgcc cgccttctgt atattagaga aaaaaagaga gcatgtgcgt gtgtgtgtat 60 ctgtgtgtct gtgtgagaga tggagaaaga gatacccttc aaaaggtgga gctggagcag 120 aatcacgact tcggaagttc agcaatgtag gctgatgcac aaatctcagg acttacgtaa 180 cagaggaaga agtgatgcca tttgtaataa aacttacctt gtctaccagc cagtggtcag 240 ggtgaggcag ccctctgggg cctactcacc catctccctt agacttgaca caactgaggc 300 agagttgcat atgcttatag tttgcttaga caattatttc tttcccatag atgatgcaaa 360 ggtatacctt tgttcagtcc taaacattag gtttttgctg gattcatgct tggggcaagg 420 gatgcatgca tacgttgatt tatttttaat ttctgtgatc atttttcatt tatttcgttg 480 ccaaagacat gtcttgctcc cattctgtca taggcatctt taaggggaaa acagcccaac 540 cctgaaagac ttaaaaggca gataagkctt ccctctttat ctggccgagg gaaattagcc 600 tgggtaagag ttccaatgag ctagcagagc ttcggctctc tccagaagtc aacaaccagt 660 tcagccaaac tgaccaacat tcctttctcc cacctgaaaa gtaaaccatc aaatctcaca 720 gatgaccgga ttgcagtgtc attattgaaa aagttgggcc tgactctccg ggaaggcacc 780 aataactagt ttaaatgtac caaggtttt 809 <210> 15 <211> 601 <212> DNA <213> Homo sapiens <400> 15 ctactcggga ggctgaggca tgagaatcac ttgaactctg gaggcagagg ttatagtgag 60 ctgagatcat gccactgcac tccagcctgg gtgatagagt gagactctgt ctcattaaaa 120 aaaaaaaaaa aattggtctc atctttcctt agttgaaagt gaaattttac ctctctgaac 180 aagtgttttc ccaagtgaaa taaatacagg aagaataaaa tctgctgata atgaaaaaaa 240 ggtggaagtc attctaagaa gtgggcagaa taaatgtgca tcttacttcg acagtcctgc 300 yggagggcat caccatagta tccaaaccgg cagaactgac agtgttcccc ttccgtgtgg 360 tacaggcact tgagacacct cccagtctcc ttgtcacagg cttctgggtc tgtcgtgtca 420 atgttgttgt gacactggca aggctgacac gaccccccaa cttctgatgg attgccaaag 480 tatcctgagg cacagtcgtc acatctggaa cctgtcaccc gataaaacca aacacaagat 540 gtttagctta ttgactgaaa gctcaccagt gcaccgacac tcatgcctag agagagctga 600 t 601 <210> 16 <211> 1280 <212> DNA <213> Homo sapiens <400> 16 ttggggatga atagatgaaa aaatatcctt gcctcagtat ttaccatgct aggttgataa 60 ctttggagca taagtagaga gaagaaaagt ttcttgtaaa gacacatttc ttaaaccaac 120 caaaagcctt atcagtgaat gctccaaagc caatttgtga gtkatagaat tacttagctt 180 taaatatgac tcttctcagg aaggcatggt cctgatcctt attttgtgct cactgacaaa 240 gctataaaaa tgcttctgaa atggggtctt tacaaagcat gtatgttcca ctacacccca 300 aaacactgct ttaccgacct gccacacact cccctactca caggccccaa gttggcttcc 360 tccgcttcat agaggtagtg atccagggtg gcaaagtagt aaccaggttc cacttcgttg 420 cactgacgtc caatcatgtg aggccggcat gagcactggc ctgactccgc aaagcaactg 480 gaagggagga ggagccacat cagctgagtt catggtcacg cgagtcaacc cgccagaagc 540 agctctacag ctgccattac tcagtacaca ggaccaaata catagcatgt tcttgttttc 600 ttcagaaaat acatatgtgg aaaaaagtag tgcctatgga ttaatcccaa actccagaag 660 cgtggctgag aacctggtct tctcggtttt acctcccttg ccctctcttg actaacacgc 720 cgtccgcatc acatgaaggt tttccacatc ccaaaccttc ttgctttggc atgttacatc 780 ttttgcttat ttcctgagat ttaggctcaa atatccagct ccctgaagaa catctccagc 840 ttggctgtcc cagagcatcc caaaccaacg gatctaagac agaagccctc aatccctcgg 900 atggtctcta tcagggtgaa ggcactacca cctgctgagt caaccaagtc agaaatttag 960 tccccgttct tctccagatt ctttcttcac actccatcag gaaccaaatt ctgtgatttt 1020 tacttctgca tcttgtgtct ctcttccccc tttccattgg ctatgctatg ctaaatataa 1080 gaaatgcggc tgggcacggt ggctcacgcc tgtaatctca gcactttggg aggccgagac 1140 aggcggatca cctgaggttg ggagttcgag accagcctca ccaacatgga gaaaccccat 1200 ctctactaaa aatacaaaat tagctgggcg tggtggcgca ggcctgtaat cccagatact 1260 tgggaggctg aggcacgaga 1280 <210> 17 <211> 601 <212> DNA <213> Homo sapiens <400> 17 aacaatcatt tgtggcatgt gttaacaaat ataatgaaat gtcaagtttg ccaattcctc 60 tggaagtatc attttcaatt tatgattaca gtgtcactta atgctgatgt accctggaaa 120 gtgggtgtca gggtagaaaa aagaaaaaag gtgagaatgt atgcaagctg tttaacagtg 180 tgcccttcat tactcccctc agaaaggccc tgttcttcat tatcaaaaaa ctcatatcac 240 ctctgtcata tttaccacac atcttttgtc ataaatagca tccaattagc aaaattttta 300 ygcagggcag cacataaaaa gatttctgcc atacatttaa atggtattca caagaggtaa 360 tctgtggtcc tttttacttt atacagtagg ctatacaacc aagcaagaaa actaattcta 420 attgttctgt ttctctataa agtaactgct gagtggctaa atagagaata ttcagctttt 480 gtttataatt tagtaacacc tgtcttttta aatcccaaat acacttctta agataaacct 540 ttttttaatg gggagaaaat gatgtgtaca gcattgattg atctttcttt gtggatttgt 600 t 601 <210> 18 <211> 977 <212> DNA <213> Homo sapiens <400> 18 gagcgcctag tagaaaatta gaagattgtg acttagcgga gtcatattca tgtccattca 60 ttgaagttca ttgacagcaa aaagccaggc ctccggatgt tcatttaaat taagtagtca 120 atatcctaat tcagtgactc gcagagaatc tggtgggtta tttaatcatc acataaaata 180 ggtggttcat gtactcatgt tgaggcagga gactaggttc tggaggcagg gaaactaagg 240 acttccttga actacattga aaggaaaacc ctaactttcc acgcctaagt aacaaaagga 300 ccagaggcta cctttgacct tttctgccta gtagatggga aattggctgt ctgtaaccaa 360 tcagactgac tgcgggtcca atcttcattt gcaactttgt aacttcactc cagcctctga 420 atggttgttg tccacaacta atcagactgt gggcgagtct tcgtttgcat agaagcrtaa 480 cttcattaag tacatcccag cctctgattg gttgtttttt gcaaccaatc agatgtttgc 540 acaggagtgt tacctttgta tcttcatttc agcctctggt tggctgcttt ctgcaaccag 600 tcagactgat tgcgggctac cacttcattt acgtgagatg agcatgagtt gccaaaggga 660 aacttcttgg gggtatttgg acccaagaag attctgtatt cggcctttga gccgctgctc 720 cgcccgctcc cacaatgtag agtgtacttt cgttttcaat aaatccctgc tttcgttctt 780 ttgtcgcttc tctgggtgtt ttgtccaatt ctttgttcaa aacgccaaga acctggacaa 840 cctgcagtca cgatccttta ctggtgacaa tattgtttta ttttactgag attccatgta 900 tcagtgttag acacaaaaat catttggtca tgtgctgcca taaataacta taaagaaagt 960 ctatgtgtgc caatttc 977 <210> 19 <211> 601 <212> DNA <213> Homo sapiens <400> 19 gtcctacact atctcaaaca ctcaagagtg tggtcctatt tgtgatggag cattgggcaa 60 tgcctgatat gtgcaccaat cccttcgcct ctcttggtct caatttcccc aaactgaagg 120 aagctgagat gtctcctaag atcctttcca atttgaaaca cagcctggat tgatgatttc 180 acagttcatc cagcaggagg aagggctgat ggctgtggga cccccatgaa agtgccttcg 240 cactctcatg aaggaattca caccaacctt cagggtaact gaagactgag cagctgccca 300 ygtgcctggc cttcttcaca ccaagctcag agcagctact gtcattccac tgttcccacc 360 atccgaatcc atagaataaa aggaggaaag agtcactttg gaaaagttag acaacaattt 420 gggaatgagg gaaaggggat gaattctgac atttttaaga gggctggagg caaatgttaa 480 ctatggcaaa gtactccagt aggttgtttt gtacgagtct cagtctaaat gaccttactg 540 tatggtatta aagaatgagg aagaaccaat aatttgatac cagatgcatg taatgaaata 600 t 601 <210> 20 <211> 509 <212> DNA <213> Homo sapiens <400> 20 gtgttctctc atgcccagtg gtgtggccta ctttgcagaa gctcatgaat gccgggattg 60 tgtcccacca gctgattaaa aacaagggaa catgttttta atgattaaac atgattaaaa 120 atatgtcagc caatagcaat gggtggatct tacttagata ctaatatgtg aagttctaac 180 ctgtacacct taagtccctc atttcttcat tgcacaaata ttgattgaga tcctcctatg 240 tgccagacrc tcttataatc actgggaacg caggattgag gaaaacagac tcagtcctta 300 tcctcgtggt gtcatcgact tcatggtcct tgaagcagta gccaggcact gggacacagt 360 ctaggtgctg gagaaccctc acactggggc aggaaagtgg tggtgtttag gtttcagtcc 420 ttgttgcatt gctctgttat aaggtcttat cttatatatg tcattttgac aaggtaaaga 480 ataagttatt catctccttc tgctttgac 509

Claims (11)

In a polynucleotide consisting of the DNA sequence of SEQ ID NO: 17, a polynucleotide consisting of 10 or more consecutive DNA sequences comprising the 301th base (polymorphic site) is T and the complementary nucleotide thereof. The polynucleotide of claim 1 or a complementary polynucleotide thereof, wherein the polynucleotide is 10 to 100 nucleotides in length. Amplification primers for diagnosing autism in Koreans, comprising the polynucleotide according to claim 1 or the complementary polynucleotide thereof. A probe for diagnosing autism in Korean, comprising the polynucleotide according to claim 1 or the complementary polynucleotide thereof. Microarray comprising a probe for diagnosing autism in Korean according to claim 4. A kit for diagnosing autism in Koreans, comprising the polynucleotide according to claim 1 or the complementary polynucleotide thereof. In order to provide information necessary for diagnosing autism in Koreans, (i) obtaining a nucleic acid sample from a sample; And (ii) analyzing the nucleotide sequence of the polymorphic site of the polynucleotide of SEQ ID NO: 17 or its complementary nucleotide from the nucleic acid sample, wherein the polymorphic site refers to the 301th base of the DNA sequence of SEQ ID NO. A method for detecting a monobasic polymorph in a sample comprising a. 8. The method of claim 7, wherein analyzing the nucleotide sequence of the polymorphic site comprises a polynucleotide of SEQ ID NO: 17 or a complementary polynucleotide sequence thereof comprising a polymorphic site, wherein the polymorphic site is a DNA sequence of SEQ ID NO: 17 Method of detecting a single nucleotide polymorphism in a sample, characterized in that carried out using a primer or a probe with a template). 8. The method of claim 7, wherein analyzing the nucleotide sequence of the polymorphic site comprises hybridizing the nucleic acid sample to a microarray to which the polynucleotide of claim 1 or a complementary nucleotide thereof is immobilized; And analyzing the obtained hybridization result. 10. The method of claim 9, wherein the DNA sequence of the polynucleotide or complementary nucleotide thereof immobilized on the microarray is 10 to 100 nucleotides in length. The method of claim 7, wherein analyzing the nucleotide sequence of the polymorphic site comprises analyzing the co-dominant genotype and the dominant genotype of the polymorphic site of SEQ ID NO: 17. 9.

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