KR101138866B1 - Genetic polymorphisms associated with myocardial infarction and uses thereof - Google Patents

Abstract

The present invention relates to gene polymorphisms associated with myocardial infarction. More specifically, the present invention includes a polynucleotide comprising a single nucleotide polymorphism associated with myocardial infarction or a complementary polynucleotide thereof, a polynucleotide hybridizing thereto, a polypeptide encoded by the antibody, an antibody binding to the polypeptide, and the polynucleotide. The present invention relates to a microarray and a kit, a diagnostic method for myocardial infarction, a monobasic polymorphism detection method, and a method for screening a medicament for treating myocardial infarction.

Description

Genetic polymorphisms associated with myocardial infarction and uses according to myocardial infarction

The present invention relates to gene polymorphisms associated with myocardial infarction and uses thereof.

99.9% of human genome sequences are identical. However, a partial 0.1% difference in the human population can lead to differences in appearance, behavior, and disease susceptibility among individuals. In other words, the differences between individuals, or between groups, races, and ethnic groups, may occur due to differences in about three million sequences. Differences in disease distribution can be associated with differences in phenotypes such as skin color among races. Of the 3 billion human genome sequences, approximately 1.0 kb may come from different bases. This totals 3 million and is called Single Nucleotide Polymorphism (SNP). In other words, if you analyze the 3 million nucleotide sequences showing polymorphism without analyzing the entire nucleotide sequence, you will find the difference between individuals or groups, or between the disease group and normal people.

The primary goal of genetics is to link differences in phenotypes such as human diseases with differences in DNA. The best tool to achieve this goal is a polymorphic marker that exists in all genomes. To date, the diversity marker used to distinguish individuals or to find related genes in genetically-differentiated families has been a microsatellite marker, but since the development of the DNA chip, interest in single nucleotide polymorphism has increased. Single nucleotide polymorphism is high in frequency, stable, and evenly distributed throughout the genome. SNP combines DNA chip technology, high-speed sequencing technology, and advanced biotechnology to advance into the new field of predictive medicine in the 21st century by identifying individual disease predictions and individual differences in medicine and revolutionizing treatment. Will contribute.

Monobasic polymorphism is an analysis of genotypes distributed at a constant frequency within a population. Therefore, the population can be classified according to genotype, and when the disease group is significantly distributed according to the genotype, the genotype and the disease can be related. For most monobasic polymorphism studies, if a single genotype or several genotypes have significantly different distributions in a disease group and a control group, it can be analyzed that the disease frequency varies depending on the genotype.

Of the more than 3 million SNPs in the human genome, about 1/6 of the 510,000 mutations are in the gene. It is very important to know their distribution because mutations in these genes are directly linked to the amount of gene expression or the function of the protein. If a genotype that is thought to be associated with a disease can cause a change in the expression or function of a gene, the gene or protein is likely the cause of the disease. In this case, the gene is a good target gene for disease treatment. Of course, disease susceptibility can also be analyzed by analyzing the corresponding monobasic polymorphism.

SNPs in a transcriptional regulatory sequence, such as a promoter, can regulate the amount of gene expressed. Very rarely, sequences in exon-intron boundaries that affect RNA splicing or, in some cases, 3'-UTR, affect RNA stability or translation efficiency. SNPs associated with disease can be found even when monobasic polymorphisms are present at the encoding site or transcriptional and translational regulatory sites and do not directly affect the protein. That is, it can be used as a useful index for determining disease susceptibility. In this case, it has not been found yet, but it may be directly affected by gene expression or protein, or it may be inherited in association with such a nucleotide sequence.
Haplotypes have more information than SNPs and contain linkage disequilibrium information, so genetic analysis by Haplotypes is more powerful. In particular, in the case of genes in which several SNPs are concentrated in the gene, genetic analysis using a haplotype combining information of adjacent SNPs is effective to reveal the association between gene function and disease. For example, in the case of Five Lipo-oxygenase Activation Peptides (FLAPs), haplotypes were found to be more potent markers for the prognosis of genetic predisposition to heart failure than SNPs. al., Nat Genet. 2004 Mar; 36 (3): 233-9). Based on these findings, deCode genetics, which conducted this study, is developing a new drug for heart attack, which is currently in Phase II clinical trials.

On the other hand, cardiovascular disease is a leading cause of death in industrialized countries around the world, and has been a leading cause of death in Korea since the 1970s. According to the National Statistical Office, 22,000 people (9087 deaths per 100,000 people), 9.1% of all Korean deaths (246,000) during 2003, died of heart disease and hypertension, leading to cancer (first place) and cerebrovascular disease ( Second place) followed by third place of death cause.

Coronary artery disease, which is an important part of cardiovascular disease, is usually caused by blockage or narrowing of coronary arteries that supply blood to the heart by arteriosclerosis. The complete blockage of coronary arteries by atherosclerosis is called angina pectoris and narrowing. The causes of coronary artery disease are hyperlipidemia (hypercholesterolemia), hypertension, smoking, diabetes, heredity, obesity, lack of exercise, stress and menopause of women. The combination of risk factors increases the risk of illness.

The biggest problem in the diagnosis or early prediction of conventional cardiovascular diseases, especially complex diseases including myocardial infarction, is that these diseases must be advanced to some extent to be diagnosed using physical methods. Currently, cardiovascular disease is diagnosed by X-ray and ultrasound imaging of the inside of the heart and coronary artery using an imaging device, but this can only be diagnosed after onset.

The present inventors continued the study to discover SNPs related to myocardial infarction, and thus, the present invention was completed by identifying SNPs related to myocardial infarction that can predict the incidence probability and genetic sensitivity of myocardial infarction.

It is an object of the present invention to provide an SNP associated with myocardial infarction.

Another object of the present invention is to provide a polynucleotide that specifically hybridizes with the polynucleotide comprising the SNP or its complementary polynucleotide.

Another object of the present invention is to provide a polypeptide encoded by the polynucleotide.

Another object of the present invention is to provide an antibody that specifically binds to the polypeptide.

Still another object of the present invention is to provide a microarray for detecting monobasic polymorphism comprising the polynucleotide.

Still another object of the present invention is to provide a kit for detecting monobasic polymorphism comprising the polynucleotide.

It is another object of the present invention to provide a method for identifying a subject having an altered risk of developing myocardial infarction.

Another object of the present invention is to provide a method for detecting monobasic polymorphism (SNP) in a nucleic acid molecule.

Still another object of the present invention is to provide a method for screening a medicament for treating myocardial infarction.

Another object of the present invention is to provide a method for regulating gene expression.

In order to achieve the object of the present invention, the present invention is a polynucleotide selected from the group consisting of SEQ ID NOs: 1 to 60, a polynucleotide comprising 8 or more consecutive nucleotides each containing the 101 st base or a complementary poly Provide nucleotides.

In order to achieve another object of the present invention, the present invention provides a polynucleotide that specifically hybridizes with the polynucleotide or its complementary polynucleotide.

In order to achieve another object of the present invention, the present invention provides a polypeptide encoded by the polynucleotide.

In order to achieve another object of the present invention, the present invention provides an antibody that specifically binds to the polypeptide.

In order to achieve another object of the present invention, the present invention provides a microarray for detecting monobasic polymorphism comprising the polynucleotide, a polypeptide encoded by the same or cDNA thereof.

In order to achieve another object of the present invention, the present invention provides a kit for detecting monobasic polymorphism comprising the polynucleotide, a polypeptide encoded by the same or cDNA thereof.

In order to achieve another object of the present invention, the present invention comprises the steps of: a) separating the nucleic acid sample from the diagnostic subject; And b) determining an allele of the polymorphic site, each 101 base of one or more polynucleotides selected from the group consisting of SEQ ID NOs: 1 to 60; and a method having a changed risk of developing a myocardial infarction. To provide.

In order to achieve another object of the present invention, the present invention is a) a polynucleotide selected from the group consisting of SEQ ID NOs: 1 to 60, a polynucleoside comprising at least 8 contiguous nucleotides each comprising the 101 st base Contacting a test sample comprising nucleic acid molecules with a reagent that specifically hybridizes under stringent hybridization conditions with the teat or its complementary polynucleotides; And b) detecting the formation of hybridized double strands; and a single nucleotide polymorphism (SNP) within a nucleic acid molecule.

In order to achieve another object of the present invention, the present invention is a) a polynucleotide selected from the group consisting of SEQ ID NOs: 1 to 60, a polynucleoside comprising at least 8 contiguous nucleotides each comprising the 101 st base Contacting the polypeptide encoded by the tit or its complementary polynucleotide with a candidate substance under suitable conditions of binding complex formation; And b) detecting the formation of a binding complex between the polypeptide and the candidate substance. It provides a method for screening a medicament for treating myocardial infarction.

In order to achieve another object of the present invention, in a polynucleotide selected from the group consisting of SEQ ID NOs: 1 to 60, a polynucleotide comprising 8 or more consecutive nucleotides each including the 101 st base, or a complement thereof It provides a method for regulating gene expression comprising binding an antisense nucleotide or Si RNA specific for a polynucleotide with the polynucleotide.

In order to achieve another object of the present invention, in a polynucleotide selected from the group consisting of SEQ ID NOs: 1 to 60, a polynucleotide comprising 8 or more consecutive nucleotides each including the 101 st base, or a complement thereof It provides a method for regulating gene expression comprising binding an antisense nucleotide or Si RNA specific for a polynucleotide with the polynucleotide.

Hereinafter, the present invention will be described in more detail.

One aspect of the invention relates to SNPs associated with myocardial infarction.

Myocardial infarction-related SNPs according to the present invention, in the polynucleotide selected from the group consisting of SEQ ID NOs: 1 to 60, comprises a polynucleotide comprising 8 or more contiguous nucleotides comprising each 101th base or a complementary polynucleotide thereof Characterized in that.

The polynucleotide comprising the polynucleotide sequence of SEQ ID NOS: 1 to 60 is a polymorphic sequence. A polymorphic sequence refers to a sequence comprising a polymorphic site representing SNP in a polynucleotide sequence. The polynucleotide can be DNA or RNA.

In the present invention, SNP (single nucleotide polymorphism) is the most present form of DNA sequence polymorphism (polymorphism) due to the difference in single base-pair variation in the DNA between the individual and the individual (about 1 / 1kb).

In one embodiment of the present invention, a series of screening procedures were performed to develop SNPs that are highly related to cardiovascular disease, especially myocardial infarction. In other words, DNA is isolated and amplified from the blood of myocardial infarction patients and normal people, and the sequence of the SNP site in the DNA is analyzed. And their genotypes. The 60 SNPs and their genotypes identified in the examples of the present invention are shown in Tables 1 and 2.

TABLE 1

SEQ ID NO: alias_id cas_num con_num Allele A Allele a cas_AA cas_Aa cas_aa con_AA con_Aa con_aa GTX p-val allele_OR One MI_0042 213 184 A G 2 44 167 10 49 125 0.00777 0.550 2 MI_0050 220 190 T G 8 57 155 2 35 153 0.0353 1.74 3 MI_0056 218 185 T G 7 58 153 One 31 153 0.00669 2.02 4 MI_0070 220 187 A G 10 54 156 9 68 110 0.0301 0.677 5 MI_0100 216 189 C T 159 54 3 124 55 10 0.0397 1.53 6 MI_0127 217 190 C T 127 79 11 126 62 2 0.0347 0.693 7 MI_0159 221 191 A G 2 39 180 2 17 172 0.0221 1.85 8 MI_0177 218 190 A G 0 12 206 0 22 168 0.0312 0.461 9 MI_0232 215 189 C T 43 109 63 53 98 38 0.0450 0.708 10 MI_0235 216 189 A G 5 63 148 2 33 154 0.00865 1.87 11 MI_0292 212 189 T G 16 102 94 5 64 120 0.000255 1.90 12 MI_0294 221 191 A G 138 77 6 100 76 15 0.02 1.52 13 MI_0299 220 191 A G 36 110 74 61 85 45 0.000629 0.596 14 MI_0354 222 190 A G 98 101 23 62 96 32 0.027 1.47 15 MI_0370 222 191 A G 44 119 59 73 83 35 0.000155 0.584 16 MI_0374 219 187 A G 5 48 166 One 25 161 0.023 1.96 17 MI_0393 222 189 C T 50 125 47 31 82 76 0.000151 1.67 18 MI_0433 222 191 A G 61 106 55 72 86 33 0.0444 0.698 19 MI_0464 222 189 A G 27 95 100 31 96 62 0.0363 0.703 20 MI_0493 221 191 A G 85 102 34 61 82 48 0.0422 1.40

Table 1 (continued)

SEQ ID NO: allele_OR_LB allele_OR_UB con_HWX_p-val gene_name SNP_function AA_change AA_position One 0.369 0.82 0.0918 intergenic intergenic n / a n / a 2 1.15 2.64 1.00 LOC144678 intron null null 3 1.30 3.13 0.641 LOC144678 mrna-utr null null 4 0.479 0.958 0.810 FLJ11117 mrna-utr null null 5 1.06 2.23 0.249 intergenic intergenic n / a n / a 6 0.49 0.98 0.0402 intergenic intergenic n / a n / a 7 1.08 3.18 0.0988 intergenic intergenic n / a n / a 8 0.225 0.944 0.476 intergenic intergenic n / a n / a 9 0.536 0.934 0.665 DUSP10 locus-region null null 10 1.23 2.86 0.392 KIAA1573 mrna-utr null null 11 1.37 2.63 0.241 DSCR1 intron null null 12 1.10 2.10 One intergenic intergenic n / a n / a 13 0.452 0.786 0.144 intergenic intergenic n / a n / a 14 1.11 1.95 0.662 KIAA1363 intron null null 15 0.442 0.77 0.176 MANBA mrna-utr null null 16 1.21 3.17 0.984 PAPSS1 coding-synon K 12 17 1.26 2.21 0.281 MANBA intron null null 18 0.529 0.92 0.45 intergenic intergenic n / a n / a 19 0.529 0.934 0.656 intergenic intergenic n / a n / a 20 1.06 1.84 0.0588 FLJ40288 mrna-utr null null

Table 1 (continued)

SEQ ID NO: alias_id cas_num con_num Allele A Allele a cas_AA cas_Aa cas_aa con_AA con_Aa con_aa GTX p-val allele_OR 21 MI_0495 221 191 A G 136 68 17 90 84 17 0.011 1.49 22 MI_0507 222 191 T G 8 60 154 8 72 111 0.0498 0.69 23 MI_0526 221 191 C A 23 97 101 33 91 67 0.0375 0.685 24 MI_0577 215 185 A G 139 64 12 134 50 One 0.00782 0.636 25 MI_0606 222 190 C G 142 68 12 136 53 One 0.00809 0.647 26 MI_0720 221 190 A G 12 73 136 21 73 96 0.0299 0.648 27 MI_1005 221 189 C A 16 66 139 12 92 85 0.000418 0.643 28 MI_1022 221 190 A G 5 62 154 2 35 153 0.0397 1.7 29 MI_1028 220 190 T G 193 27 0 181 9 0 0.00821 0.371 30 MI_1029 221 190 C T 46 124 51 60 92 38 0.0468 0.757 31 MI_1036 219 191 C T 14 102 103 29 90 72 0.00832 0.667 32 MI_1039 217 189 A G 154 59 4 159 27 3 0.00415 0.524 33 MI_1051 222 191 A G 131 85 6 95 80 16 0.0172 1.48 34 MI_1065 219 189 A G One 36 182 One 50 138 0.0193 0.596 35 MI_1070 216 185 T G 28 107 81 41 93 51 0.0189 0.675 36 MI_1071 222 190 T G 2 39 181 4 51 135 0.0384 0.583 37 MI_1076 215 190 T G 97 96 22 108 72 10 0.0303 0.662 38 MI_1096 221 191 A G 0 6 215 0 15 176 0.0235 0.337 39 MI_1112 222 191 A G 115 94 13 123 62 6 0.0283 0.649 40 MI_1130 222 190 C G 155 64 3 150 40 0 0.0359 0.629

Table 1 (continued)

SEQ ID NO: allele_OR_LB allele_OR_UB con_HWX_p-val gene_name SNP_function AA_change AA_position 21 1.09 2.03 0.741 intergenic intergenic n / a n / a 22 0.49 0.972 0.537 GNA12 intron null null 23 0.515 0.911 0.775 intergenic intergenic n / a n / a 24 0.437 0.925 0.135 ALOX5AP intron null null 25 0.449 0.934 0.0795 ALOX5AP intron null null 26 0.473 0.887 0.165 LGALS2 intron null null 27 0.470 0.88 0.0571 intergenic intergenic n / a n / a 28 1.12 2.58 One ANK3 mrna-utr null null 29 0.172 0.799 One HIP1 intron null null 30 0.575 0.997 0.776 intergenic intergenic n / a n / a 31 0.499 0.892 0.883 intergenic intergenic n / a n / a 32 0.337 0.815 0.15 intergenic intergenic n / a n / a 33 1.08 2.03 0.865 intergenic intergenic n / a n / a 34 0.382 0.928 0.212 intergenic intergenic n / a n / a 35 0.509 0.895 One THH intron null null 36 0.384 0.888 One MAP2K4 intron null null 37 0.486 0.902 0.692 intergenic intergenic n / a n / a 38 0.129 0.877 One intergenic intergenic n / a n / a 39 0.467 0.901 0.815 RGS7 intron null null 40 0.415 0.952 0.23 RBL2 mrna-utr null null

Table 1 (continued)

SEQ ID NO: alias_id cas_num con_num Allele A Allele a cas_AA cas_Aa cas_aa con_AA con_Aa con_aa GTX p-val allele_OR 41 MI_1145 221 191 A G 23 99 99 34 93 64 0.0217 0.67 42 MI_1169 212 186 A G 3 37 172 0 19 167 0.0207 2.10 43 MI_1175 222 187 A G 0 26 196 One 36 150 0.0315 0.55 44 MI_1186 219 188 A G 149 64 6 97 73 18 0.000435 1.94 45 MI_1206 222 190 A G 80 113 29 57 90 43 0.035 1.38 46 MI_1209 218 190 T G 16 77 125 16 89 85 0.0374 0.713 47 MI_1221 219 190 A G 2 40 177 0 18 172 0.00955 2.25 48 MI_1247 215 186 C T 54 101 60 67 85 34 0.0193 0.661 49 MI_1261 222 189 C T 171 50 One 165 22 2 0.00513 0.557 50 MI_1264 222 190 C T 14 55 153 2 46 142 0.0182 1.52 51 MI_1272 221 188 A G 26 106 89 38 91 59 0.0332 0.696 52 MI_1273 221 188 C A 188 30 3 136 45 7 0.00547 2.10 53 MI_1329 220 189 C G 71 115 34 93 74 22 0.00255 0.637 54 MI_1363 221 190 T A 103 102 16 67 97 26 0.0199 1.48 55 MI_1377 222 190 C T 86 108 28 100 72 18 0.0181 0.678 56 MI_1503 216 187 A G 0 30 186 2 42 143 0.0153 0.532

Table 1 (continued)

SEQ ID NO: allele_OR_LB allele_OR_UB con_HWX_p-val gene_name SNP_function AA_change AA_position 41 0.504 0.89 One intergenic intergenic n / a n / a 42 1.20 3.67 One SIPA1L1 intron null null 43 0.327 0.924 0.413 intergenic intergenic n / a n / a 44 1.39 2.71 0.38 intergenic intergenic n / a n / a 45 1.04 1.82 0.476 CSMD1 intron null null 46 0.525 0.969 0.316 intergenic intergenic n / a n / a 47 1.27 3.96 One LOC387895 intron null null 48 0.499 0.874 0.449 intergenic intergenic n / a n / a 49 0.34 0.911 0.212 intergenic intergenic n / a n / a 50 1.04 2.22 0.746 MST1R coding-nonsynon G 1335 51 0.525 0.923 0.769 SLC8A1 intron null null 52 1.35 3.26 0.0919 intergenic intergenic n / a n / a 53 0.478 0.851 0.168 NFKB1 intron null null 54 1.11 1.98 0.361 intergenic intergenic n / a n / a 55 0.505 0.91 0.369 NFKB1 intron null null 56 0.328 0.862 One CYBA coding-nonsynon H 72

In Tables 1 and 2, the column 'SEQ ID NO' refers to the number of the polynucleotide sequence comprising each SNP attached to this specification.

The column 'alias_id' means a number of SNPs named by the present inventors arbitrarily.

The columns 'cas_num' and 'con_num' represent the number of patient groups and normal human groups each having a corresponding SNP site.

Allele A and allele a are randomly named alleles of small mass and alleles of a for convenience of experiments during sequencing by Sequenom's homogeneous MassEXTEND technique. .

Columns 'cas_AA', 'cas_Aa' and 'cas_aa' represent the number of patients with genotypes AA, Aa and aa, respectively, and columns 'con_AA', 'con_Aa' and 'con_aa' are genotypes of AA, Aa and aa, respectively The number of normal persons having

The column 'GTX_p-val' refers to a p-value that is the result of Fisher's exact test for the genotype. If p-value ≤ 0.05, the genotypes of the disease group and the normal group were not the same.

The column 'allele_OR' represents the odds ratio, which is the probability of having the SNP among the patient group with respect to the probability of having the SNP among the normal group based on the genotype. The columns 'allele_OR_LB' and 'allele_OR_UB' mean lower and upper bounds of the 95% confidence interval for the probability that the corresponding odds ratio exists in a certain interval. When the odds ratio is greater than 1, A is a risk factor of myocardial infarction, and if it is less than 1, a is a risk factor of myocardial infarction. In addition, when the confidence interval includes 1, the association with the disease in the experimental group irradiated with the genotype cannot be determined to be significant.

The column 'con_HWX_p-val' represents the Hardy-Wineberg equilibrium in the normal group. The p-value of 0.05 or less in the assay was determined by Hardy-Weinberg Equilibrium.

The column 'gene_name' is the name of the gene to which the SNP belongs.

The column 'SNP_function' means the role played by each single SNP in the gene.

Column 'AA_change' indicates whether the amino acid is changed by the SNP.

Column 'AA_position' indicates the position on the polypeptide of the amino acid encoded by the SNP site.

TABLE 2

SEQ ID NO: alias_id Allele A Allele a gene_name SNP_function AA_change AA_position cas_a 57 MI_1111 A G polymerase iota intron null null 0.287 58 MI_1248 C T polymerase iota exon Thr-> Ala 706 0.715 59 MI_2143 T G polymerase iota intron null null 0.285 60 MI_2144 T G polymerase iota intron null null 0.322

Table 2 (continued)

SEQ ID NO: con_a Delta chi_value chi_exact_Pvalue OR OR_LB OR_UB con_HW 57 0.215 0.072 7.561 0.0228 1.47 1.113 1.937 HWE 58 0.788 0.075 8.043 0.0179 1.5 1.131 1.978 HWE 59 0.217 0.068 6.829 0.0329 1.49 1.094 1.904 HWE 60 0.242 0.8 8.816 0.0122 1.49 1.137 1.949 HWE

The columns 'cas_a', 'con_a' and 'Delta' represent the frequency of the allele in the patient group, the frequency of the allele in the normal group, and the absolute value of the difference between the cas_a and con_a, respectively. Where cas_a is (frequency of aaa genotype × 2 + frequency of Aa genotype) / (number of samples in disease group × 2) in the patient group and con_a is (frequency of aaa genotype × 2 + frequency of Aa genotype) / (normal in normal group) Number of samples in the group x 2).

The column 'Chi-value' is the value of the chi-square test and is the value for the p-value calculation. The column 'chi-exact-p-value' represents the p-value of Fisher's exact test of chi-square test, and if the value of the genotype number contains a value less than 5, the general chi-square test results may be inaccurate. This is a variable used to test more accurate statistical significance (p-value) through Fisher's exact test. If p-value ≤ 0.05, the genotypes of the disease group and the normal group were not the same.

The column 'OR' is a ratio that indicates whether a specific genotype is specifically detected on one side for a disease group and a normal group. It is obtained by calculating the number of disease groups without a specific genotype) * (the number of normal groups with a specific genotype). The columns 'OR_LB' and 'OR_UB' represent the minimum and maximum values of the confidence interval of OR at the significance level of 5%, respectively.
All four SNPs in Table 3 are located in the AGT gene.

In addition, the present invention includes specific haplotypes. That is, in the myocardial infarction diagnostic polynucleotide or a complementary polynucleotide thereof, the polynucleotide is preferably composed of SEQ ID NOs: 57 to 60. Linkage disequlibrium (LD) relative to each other is shown in Table 3. As shown in Table 3, the four SNPs constituted a strong LD block.

TABLE 3

MI_2144 MI_1111 MI_2143 MI_1248 MI_2144 0 One One 0.9898 MI_1111 One 0 One 0.9951 MI_2143 One One 0 One MI_1248 0.9898 0.9951 One 0

More preferably, the 101 st base, which is the SNP region of the polynucleotides SEQ ID NOs: 57 to 60 constituting the haplotype, may be each risk allele. That is, it may be Haplotype No. 1 or 2 in Table 4.

TABLE 4

Haplotype type number MI_2144 MI_1111 MI_2143 MI_1248 Hap.score p.val Hap.Freq total_freq One One One One 2 -3.06506 0.00218 71.60% 2 2 2 2 One 2.71413 0.00664 25.00% 3 2 One One 2 0.73651 0.46142 3%

In Table 4, the number '1' or '2' of each SNP column means the number of alleles. For example, column 1 is a haplotype consisting of four SNPs and a haplotype consisting of alleles of '1', '1', '1', and '2' at each position. The column 'Hap.score' is a measure of how well a particular haplotype distinguishes between a normal group and a disease group, and is judged to be significant when the value of the column 'p.val' is 0.05 or less.

The association of habitual or environmental factors associated with myocardial infarction of the SNPs of SEQ ID NOS: 57-60 was investigated. The results are shown in Table 5. As shown in Table 5, the male and non-smokers showed a more significant relationship with the development of myocardial infarction. This can be seen from the increase in the odds ratio for male and non-smokers in Table 5.

TABLE 5

group alias_id SEQ ID NO: OR OR_LB OR_UB Chi_squ_Pvalue male MI_1111 57 1.3816 1.0067 1.8961 0.0939 MI_1248 58 1.4353 1.0427 1.9757 0.0674 MI_2143 59 1.3513 0.98 1.8587 0.112 MI_2144 60 1.4705 1.08 2 0.0547 Male smoker MI_1111 57 1.8472 1.0138 3.3657 0.161 MI_1248 58 1.8752 1.029 3.4173 0.166 MI_2143 59 1.8181 1.033 3.1746 0.122 MI_2144 60 1.8518 1.0141 3.3557 0.132 Male non-smoker MI_1111 57 2.2865 1.2005 4.3546 0.0231 MI_1248 58 2.3976 1.2492 4.6018 0.024 MI_2143 59 2.2727 1.2004 4.3478 0.0242 MI_2144 60 2.5641 1.3458 4.7619 0.0107

The polynucleotide of each single SNP constituting the SNP according to the present invention is preferably 8 or more contiguous nucleotides, more preferably 8 to 70 contiguous nucleotides.

Another aspect of the present invention relates to a polynucleotide that specifically hybridizes with the polynucleotide of SEQ ID NOS: 1-60 or the complementary polynucleotide thereof according to the present invention. The polynucleotide is allele specific.

The polynucleotide is preferably 8 or more contiguous nucleotides, more preferably 8 to 70 contiguous nucleotides.

The allele-specific polynucleotide means to hybridize specifically to each allele. That is, it means hybridizing so that the base of the polymorphic site in each polymorphic sequence of SEQ ID NO: 1-60 can be distinguished specifically. Here, hybridization is usually carried out under stringent conditions, for example salt concentrations of 1 M or less and temperatures of 25 ° C. or higher. For example, conditions of 5 × SSPE (750 mM NaCl, 50 mM Na Phosphate, 5 mM EDTA, pH 7.4) and 25-30 ° C. may be suitable for allele specific probe hybridization.

In the present invention, the allele specific polynucleotide may be a primer. Here primers are the starting point for template-directed DNA synthesis under appropriate conditions (e.g. four different nucleoside triphosphates and polymerizers such as DNA, RNA polymerase or reverse transcriptase) in appropriate buffers and at appropriate temperatures. Refers to single-stranded oligonucleotides that can act. The appropriate length of the primer may vary depending on the purpose of use, but is usually 15 to 30 nucleotides. Short primer molecules generally require lower temperatures to form stable hybrids with the template. The primer sequence need not be completely complementary to the template, but should be sufficiently complementary to hybridize with the template. Preferably, the primer 3 'end is aligned with the polymorphic site of SEQ ID NO: 1 to 60. The primer hybridizes to the target DNA comprising the polymorphic site and initiates amplification of allelic forms in which the primer exhibits complete homology. This primer is used in pairs with a second primer that hybridizes to the other side. By amplification the product is amplified from two primers, which means that a specific allele form is present. Primers of the invention include polynucleotide fragments used in a ligase chain reaction (LCR). For example, the primer may be a polynucleotide of SEQ ID NO: 87 to SEQ ID NO: 344 of Table 6.

In the present invention, the allele specific polynucleotide may be a probe. Probe (probe) in the present invention means a hybridization probe, it means an oligonucleotide capable of sequence-specific binding to the complementary strand of the nucleic acid. Such probes include peptide nucleic acids described in Nielsen et al., Science 254, 1497-1500 (1991). Probes of the present invention are allele-specific probes in which polymorphic sites exist in nucleic acid fragments derived from two members of the same species, hybridizing to DNA fragments derived from one member, but not to fragments derived from other members. . In this case, the hybridization conditions show a significant difference in the hybridization strength between alleles, and should be severe enough to hybridize to only one of the alleles. In the probe of the present invention, the central region (that is, position 7 when the probe is 15 nucleotides and position 8 or 9 when the probe is 16 nucleotides) is preferably aligned with the polymorphic site of the sequence. This can lead to good hybridization differences between different allelic forms. The probe of the present invention can be used for diagnostic methods for detecting alleles. The diagnostic method includes detection methods based on hybridization of nucleic acids such as Southern blotting, or the like, and may be provided in a form that is pre-coupled to a substrate of the microarray in a method using a microarray.

Another aspect of the invention relates to a polypeptide encoded by a polynucleotide of SEQ ID NOs: 1-60.

In particular, a polypeptide encoded by a polynucleotide of SEQ ID NO: 56 comprising MI_1503 located within the gene CYBA varies in its amino acid. In addition, the polypeptide encoded by the polynucleotide of SEQ ID NO: 50 including MI_1264 located in the gene MST1R and the polynucleotide of SEQ ID NO: 58 including MI_1248 located in the gene polymerase iota changes the corresponding amino acid. The three amino acid change positions are 72th amino acid of CYBA protein, 1335th amino acid of MST1R protein and 706th amino acid of polymerase iota protein, respectively.

Another aspect of the invention relates to an antibody that specifically binds to the polypeptide. It is preferable that the said antibody is a monoclonal antibody.

Another aspect of the invention relates to a microarray comprising a polynucleotide of SEQ ID NOs: 1 to 60 according to the invention or a complementary polynucleotide thereof or a polynucleotide hybridizing with them, a polypeptide encoded by it or a cDNA thereof.

Microarrays according to the present invention are conventional methods known to those skilled in the art using polynucleotides according to the present invention or polynucleotides which hybridize with a probe or their complementary polynucleotides, polypeptides encoded by them or cDNAs thereof. It can be prepared by.

For example, the polynucleotide may be immobilized on a substrate coated with an active group selected from the group consisting of amino-silane, poly-L-lysine, and aldehyde. In addition, the substrate may be selected from the group consisting of silicon wafer, glass, quartz, metal and plastic. As a method of immobilizing the polynucleotide on a substrate, a micropipetting method using a piezoelectric method, a method using a pin type spotter, or the like can be used.

Another aspect of the present invention relates to a kit comprising a polynucleotide of SEQ ID NOs: 1 to 60 according to the present invention or a complementary polynucleotide thereof or a polynucleotide hybridizing thereto, a polypeptide encoded by the same, or a cDNA thereof.

The kit according to the present invention may further comprise a primer set used to separate and amplify DNA containing the SNP from the diagnostic subject in addition to the polynucleotide according to the present invention. Such suitable primer sets will be readily apparent to those skilled in the art with reference to the sequences of the present invention. For example, those shown in Table 6 may be used as the primer set. In addition, the kit according to the present invention may further include reagents required for the polymerization reaction, such as dNTP, polymerases and coloring agents of various species.

Another aspect of the present invention relates to a method for identifying a subject having an altered risk of developing myocardial infarction using the SNP according to the present invention.

The diagnostic method of myocardial infarction according to the present invention comprises the steps of: a) separating the nucleic acid sample from the diagnostic object; And b) determining an allele of the polymorphic site, each 101 base of one or more polynucleotides selected from the group consisting of SEQ ID NOs: 1-60.

In the method of the present invention, the method of separating DNA from a diagnosis subject can be made through a method known in the art. For example, it can be done by directly purifying DNA from tissue or cells or by specifically amplifying and isolating specific regions using amplification methods such as PCR. In the present invention, DNA is meant to include not only DNA but also cDNA synthesized from mRNA. The step of obtaining nucleic acid from the diagnostic target is, for example, PCR amplification, ligase chain reaction (LCR) (Wu and Wallace, Genomics 4, 560 (1989), Landegren et al., Science 241, 1077 (1988)), transcription amplification transcription amplification (Kwoh et al., Proc. Natl. Acad. Sci. USA 86, 1173 (1989)) and self-sustaining sequence replication (Guatelli et al., Proc. Natl. Acad. Sci. USA 87, 1874 (1990)) and Nucleic acid based sequence amplification (NASBA) can be used.

Determination of the nucleotide sequence of the isolated DNA can be made by various methods known in the art. For example, a nucleotide of a polymorphic site is determined by the dideoxy method by hybridizing the probe or complementary probes comprising the sequence of the SNP site or through the determination of the nucleotide sequence of the nucleic acid with the DNA and measuring the degree of hybridization obtained therefrom. Methods for determining sequences may be used. The degree of hybridization may be achieved by, for example, labeling a detectable label on the target DNA to specifically detect only the hybridized target DNA, but other electrical signal detection methods may be used.

Specifically, allele-specific probe hybridization, allele-specific amplification, sequencing, 5 'nuclease digestion, In the method selected from the group consisting of molecular beacon assay, oligonucleotide ligation assay, size analysis and single-stranded conformation polymorphism. Can be performed by

The method for diagnosing myocardial infarction according to the present invention c) if the allele of the polymorphic site of one or more polynucleotides selected from the group consisting of SEQ ID NOs: 1 to 60 includes a risk allele, the subject develops myocardial infarction The method may further include determining that the increased risk of P is.

In the present invention, the risk allele is A when the frequency of A in the disease group is greater than the frequency of A in the normal group, with the reference allele as A, and vice versa. Has a as the risk allele. The more risk alleles, the greater the likelihood of developing myocardial infarction.

In the method for diagnosing myocardial infarction according to the present invention, the altered risk may be increased risk or reduced risk. The altered risk may be a reduced risk if the frequency of the allele of one SNP is greater in the normal group than in the disease group, and the altered risk is increased if the frequency of the allele of one SNP is greater in the disease group than in the settlement group. May be a risk.

In the method for diagnosing myocardial infarction according to the present invention, the type of the diagnosis target is male and non-smoker, and the polynucleotide for determining the allele of the polymorphic site for the subject is selected from the group consisting of SEQ ID NOs: 57 to 60 It is preferable to be.

In addition, in the method for diagnosing myocardial infarction according to the present invention, it is preferable to use the haplotype according to the present invention. That is, the polynucleotide is preferably composed of SEQ ID NO: 57 to 60.

Another aspect of the present invention is a) a polynucleotide selected from the group consisting of SEQ ID NOs: 1 to 60, comprising a polynucleotide comprising 8 or more consecutive nucleotides comprising each 101th base or a complementary polynucleotide thereof; Contacting a reagent that specifically hybridizes under stringent hybridization conditions with a test sample comprising nucleic acid molecules; And b) detecting the formation of hybridized double strands; and a method for detecting a single nucleotide polymorphism (SNP) in a nucleic acid molecule.

Step b) includes allele-specific probe hybridization, allele-specific amplification, sequencing, 5 'nuclease digestion. ), Molecular beacon assay, oligonucleotide ligation assay, size analysis, and single-stranded conformation polymorphism. It may be carried out by the method.

Another aspect of the invention is a) a polynucleotide selected from the group consisting of SEQ ID NOs: 1 to 60, comprising a polynucleotide comprising 8 or more consecutive nucleotides comprising each 101th base or a complementary polynucleotide thereof Contacting the polypeptide encoded by the candidate substance under suitable conditions of binding complex formation; And b) detecting the formation of the binding complex between the polypeptide and the candidate substance.

Step b) may be performed by immunoprecipitation, radioimmunoassay (RIA), enzyme immunoassay (ELISA), immunohistochemistry, Western blotting or flow cytometry (FACS).

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

Example 1

Screening of SNPs According to the Invention

In this example, DNA was isolated from blood of a Korean patient who was found to be a cardiovascular disease patient and a normal patient without cardiovascular disease symptoms, and the frequency of occurrence of a specific SNP was analyzed. The SNPs selected in this example are published databases (NCBI dbSNP: http://www.ncbi.nlm.nih.gov/SNP/) or realsnp.com (http: //: //www.realsnp.com/) of Sequenom. SNP sequences in the samples were analyzed using primers using sequences around the selected SNP.

1-1. Preparation of DNA Samples

DNA was extracted from the blood of Korean male patients (221 patients) who were identified as being treated with myocardial infarction (221) and normal Korean males who had no symptoms of myocardial infarction (192). Chromosome DNA extraction was performed according to known extraction methods (Molecular cloning: A Laboratory Manual, p 392, Sambrook, Fritsch and Maniatis, 2nd edition, Cold Spring Harbor Press, 1989) and the instructions of the commercial kit (Gentra system, D-50K). Was done. Among the extracted DNA, the purity standard was selected and used only at least 1.7 in the UV measurement ratio (260 / 280nm).

1-2. Amplification of Target DNA

Target DNA, which is a constant DNA region containing the SNP to be analyzed, was amplified by PCR. PCR method proceeded in a conventional manner, the conditions were as follows. First, target genomic DNA was prepared at 2.5 ng / ml. Next, the following PCR reaction solution was prepared.

3.14 µl of water (HPLC grade)

0.5 μl 10x buffer

0.2 μl MgCl ₂ 25 mM

0.04 μl dNTP Mix (GIBCO) (25 mM / each)

Taq pol (HotStart) (5U / μl) 0.02μl

0.1 μl transposition / back end primer mix (10 μM)

1.00 μl DNA

5.00 μl total reaction volume

Here, the forward and reverse primers were selected at appropriate positions from upstream and downstream of the SNP. The primer sets are summarized in Table 6.

Thermal cycling of PCR was maintained at 95 ° C. for 15 minutes, repeated 45 seconds at 95 ° C., 30 seconds at 56 ° C., 1 minute at 72 ° C., and held at 72 ° C. for 3 minutes, followed by 4 ° C. Stored in. As a result, a target DNA fragment having a length of 200 nucleotides or less was obtained.

1-3. Analysis of SNP Site Sequences in Amplified Target DNA

The analysis of SNPs in the target DNA fragments was carried out using a homogeneous Mass Extend (hME) technique from Sequenom. The principle of the MassEXTEND technique is as follows. First, a primer (also called extension primer) complementary to the base immediately before the SNP in the target DNA fragment is prepared. Next, the primer is hybridized to the target DNA fragment, and a DNA polymerization reaction is caused. At this time, a reagent (Termination mix; for example, ddTTP) to stop the reaction after the base complementary to the first allele base (for example, A allele) among the target SNP alleles is added to the reaction solution. Include it. As a result, when a first allele (eg, A allele) is present in the target DNA fragment, a product is obtained in which only one base (eg, T) complementary to the first allele is added. . On the other hand, if a second allele (eg, G allele) is present in the target DNA fragment, the closest first allele added with a base (eg, C) complementary to the second allele A product extending to the base (eg A) is obtained. By determining the length of the product extended from the primer thus obtained by mass spectrometry, it is possible to determine the type of allele present in the target DNA. Specific experimental conditions were as follows.

First, unbound dNTPs were removed from the PCR product. To this end, 1.53 μl of pure water, 0.17 μl of hME buffer, and 0.30 μl of SAP (shrimp alkaline phosphatase) were added to a 1.5 ml tube to prepare a SAP enzyme solution. The tube was centrifuged at 5,000 rpm for 10 seconds. After that, the PCR (Polymerase Chain Reaction) product was placed in the SAP solution tube, sealed, and kept at 37 ° C. for 20 minutes and 85 ° C. for 5 minutes, and then stored at 4 ° C.

Next, a homogeneous extension reaction was performed using the target DNA product as a template. The reaction solution was as follows.

1.728 μl of water (nano-grade pure water)

0.200 μl hME extension mix (10 × buffer with 2.25 mM d / ddNTPs)

0.054 μL extension primer (100 μM each)

0.018 μl Thermosequenase (32U / μl)

Total volume 2.00 μl

The reaction solution was mixed well, followed by spin down centrifugation. Seal the tube or plate containing the reaction solution well, hold for 2 minutes at 94 ℃, then repeat 5 seconds at 94 ℃, 5 seconds at 52 ℃, 5 seconds at 72 ℃ 40 times, and then 4 ℃ Stored in. The salts were removed from the homogeneous extended reaction product thus obtained using Resin (SpectroCLEAN, Sequenom company # 10053). The extension primers used for the extension reaction are shown in Table 6.

TABLE 6

SNP-containing nucleotides
(SEQ ID NO) Target DNA Amplification Primer (SEQ ID NO :) Extension primer
(SEQ ID NO) Potential primer Back primer One 61 62 63 2 64 65 66 3 67 68 69 4 70 71 72 5 73 74 75 6 76 77 78 7 79 80 81 8 82 83 84 9 85 86 87 10 88 89 90 11 91 92 93 12 94 95 96 13 97 98 99 14 100 101 102 15 103 104 105 16 106 107 108 17 109 110 111 18 112 113 114 19 115 116 117 20 118 119 120

Table 6 (continued)

SNP-containing nucleotides
(SEQ ID NO) Target DNA Amplification Primer (SEQ ID NO :) Extension primer
(SEQ ID NO) Potential primer Back primer 21 121 122 123 22 124 125 126 23 127 128 129 24 130 131 132 25 133 134 135 26 136 137 138 27 139 140 141 28 142 143 144 29 145 146 147 30 148 149 150 31 151 152 153 32 154 155 156 33 157 158 159 34 160 161 162 35 163 164 165 36 166 167 168 37 169 170 171 38 172 173 174 39 175 176 177 40 178 179 180

Table 6 (continued)

SNP-containing nucleotides
(SEQ ID NO) Target DNA Amplification Primer (SEQ ID NO :) Extension primer
(SEQ ID NO) Potential primer Back primer 41 181 182 183 42 184 185 186 43 187 188 189 44 190 191 192 45 193 194 195 46 196 197 198 47 199 200 201 48 202 203 204 49 205 206 207 50 208 209 210 51 211 212 213 52 214 215 216 53 217 218 219 54 220 221 222 55 223 224 225 56 226 227 228 57 229 230 231 58 232 233 234 59 235 236 237 60 238 239 240

The reaction product was analyzed by the sequence Assisted Laser Desorption and Ionization-Time of Flight (MALDI-TOF) in the mass spectrometry. The MALDI-TOF is based on the principle of mass analysis by calculating the time taken to fly with an ionized matrix (3-Hydorxypicolinic acid) to fly to the opposite detector under vacuum when the material to be analyzed receives a laser beam. Works. The smaller mass reaches the detector quickly, and the SNP sequence of the target DNA can be determined based on the difference in mass and the known sequence of the SNP.

1-4. Screening of SNPs According to the Invention

We compared the frequency of alleles between the Korean male patients (221) who were diagnosed as being treated with myocardial infarction and the normal Korean male group (192) who did not have myocardial infarction. Based on the frequency, Fisher's exact test, an association test, was performed.

Effect size was estimated using the allele odds ratio and its 95% confidence interval. If the reported allele is more frequent in the normal group than in the patient group, the allele is associated with a reduced risk of myocardial infarction and the remaining allele is the risk allele of myocardial infarction. Conversely, if the reported allele appears to be higher in the patient group than in the normal group, the allele is a risk allele of myocardial infarction.

If the SNP was p-value <0.05 in the allele relevance test, the SNP was considered to be a significant genetic marker.

The results are shown in Table 1 and Table 2. As shown in Table 1 and Table 2, 60 SNPs related to myocardial infarction were identified.

1-5. Selection of haplotype according to the present invention

For the polymerase iota gene, a DNA repair-related gene, four SNPs were shown in Table 2 and linkage disequilibrium (LD) was examined. As a result, four SNPs of SEQ ID NOs: 57 to 60 showed high associative equilibrium, constituting a strong LD block. The results are shown in Table 3.

1-6. Investigation of environmental or habitual factor dependence of SNP according to the present invention

Each group of Korean male patients (221) who were identified as being treated with myocardial infarction and a normal Korean male group (192) who did not yet have myocardial infarction were divided into subgroups based on the risk of environmental or habitual factors associated with myocardial infarction. And allele frequencies were compared. That is, the association between habitual or environmental factors related to myocardial infarction of SNPs SEQ ID NOS: 57-60, in particular gender and smoking status, was investigated.

The results are shown in Table 4. As shown in Table 4, the SNPs of SEQ ID NOs: 57-60 were more significantly associated with the development of myocardial infarction in the case of males and non-smokers, respectively.

Example 2

Fabrication of SNP-Finished Microarrays According to the Present Invention

The SNPs selected above were fixed on a substrate to prepare a microarray. That is, polynucleotides consisting of 20 contiguous nucleotides each containing 101 bases in the polynucleotides consisting of SEQ ID NOs: 1 to 60 (each SNP is positioned on the 11th of the 20) were immobilized on a substrate.

First, the N-terminus of each polynucleotide was substituted with an amine group and then spotted on a silylation slide (Telechem), wherein the spotting buffer was 2 × SSC (pH 7.0). After spotting, the mixture was placed in a dryer to induce binding, followed by washing with 0.2% SDS for 2 minutes and tertiary distilled water for 2 minutes to remove unbound oligos. The slides were treated at 95 ° C. for 2 minutes to induce denaturation, followed by 15 minutes with blocking solution (1.0 g NaBH ₄ , PBS (pH 7.4) 300 mL, EtOH 100 mL), 1 minute with 0.2% SDS solution, 3rd Each was washed with distilled water for 2 minutes and dried at room temperature to prepare a microarray.

Example 3

Myocardial infarction diagnosis using microarray according to the present invention

Target DNA was isolated and fluorescently labeled from the blood of a subject to diagnose the onset or possibility of developing myocardial infarction using the method described in steps 1-1 and 1-2 of Example 1. Hybridization of the microarray prepared in Example 2 and the fluorescently labeled target DNA under UniHyb hybridization solution (TeleChem) was performed at 42 ° C. for 4 hours. It was washed twice with 2 x SSC for 5 minutes at room temperature and then dried in air. After drying, the slides were scanned with ScanArray 5000 (GSI Lumonics) and the scan results were analyzed with QuantArray (GSI Lumonics) and ImaGene software (BioDiscover) to determine the SNP according to the present invention. The presence of myocardial infarction and the susceptibility to myocardial infarction were measured by confirming whether they had.

Myocardial infarction related SNPs and haplotypes according to the present invention can be used for applications related to myocardial infarction such as diagnosis, treatment and gene fingerprint analysis of myocardial infarction. In addition, according to the microarray and kit including the SNP of the present invention, myocardial infarction can be effectively diagnosed. In addition, according to the method for analyzing the SNP associated with myocardial infarction of the present invention, it is possible to effectively diagnose the presence or risk of myocardial infarction.

<110> Samsung Electronics Co., Ltd <120> Genetic polymorphisms associated with myocardial infarction and          uses according <130> PN061465 <160> 240 <170> KopatentIn 1.71 <210> 1 <211> 200 <212> DNA <213> Homo sapiens <400> 1 aagaatggaa ggacagcttg ctacttctgg gtgttcatca tgagcagtta aggctcacca 60 tatggtctcc actgatattt gggggaagga gacttgttac yacctaataa tgagatgaaa 120 gatcctgatt ccctacttgg tttactatga caccgccctg gcatgatggt tcaggaaact 180 ctttacattt acagattgtc 200 <210> 2 <211> 200 <212> DNA <213> Homo sapiens <400> 2 tactgctgtc tttggaacac tgttactcat cctacaaggc ctcaatcaag aatcttagga 60 ggtcattggt gcctacctgc ccatggcaga gaccaccaac mccttctgag agctgccaca 120 gtgctgcttt ctaacatgtc tacactagct cctcacagtg tgcggtgatt gttgggtgaa 180 agctctgttt ctcatagtga 200 <210> 3 <211> 200 <212> DNA <213> Homo sapiens <400> 3 tgatttgatt ttatccagtg ttgcaaactg ggtttcgtaa gtgactcttt tgtaaggagt 60 ttcagtgcac cccgtctgca acccatgcat atcttattta mccccaagct tctaaggtag 120 aatcctactt ttttcaagtg ctattgatta gttgcctgtt tagacttttg ttatatgatt 180 taaaatttta aaaatgataa 200 <210> 4 <211> 200 <212> DNA <213> Homo sapiens <400> 4 gtgtttctaa agcaactccc caccaacctc ttgggctgtg gtgaccaagg gctaaaaatt 60 aagtggggac cctggtgtct gacttttgct ccacttctga yaccttcttt ctctcccgtt 120 tcactaggac tggaaaatac aggctttttt cccaatcctt gtttctctca aaccaagacc 180 tcatctacat aaaacatcta 200 <210> 5 <211> 200 <212> DNA <213> Homo sapiens <400> 5 ccactgaaaa tatcctctgg tcatttattt ggctcaacgg acctatatgg ccattccctc 60 ttccacagtt gtaaagactt tgcctaattt ctggcaacta yagaagtagt ggtaaggata 120 atgtaaaatt tctttttgtc ttggaaatta tcaaaccact tttgaagatt gtattatcta 180 gtaatcagaa acttactttg 200 <210> 6 <211> 166 <212> DNA <213> Homo sapiens <400> 6 taagggcttg agtatgcaaa tatggaggct ttctcaaatc acgtttcgtt taatttaaca 60 ccttcataaa taggatggac ttaagagatc atcccaacct ycttactgta cagatcaaga 120 gatgggggtc cggaaagctg ctgatgttca tgctgtgagt ttaaca 166 <210> 7 <211> 200 <212> DNA <213> Homo sapiens <400> 7 tttgccaacc attcttcacc ctgttgggag agaatacaga aatctgtatt caagcataag 60 gtgctataca cttgtttgat gcctgcctct agggcaatga rgaggtaaga caatctcccc 120 attacatgct ttcgcagagg acagtctgta aagttacaga actccattcc ttcttacgta 180 ccttatacca taatatacca 200 <210> 8 <211> 152 <212> DNA <213> Homo sapiens <400> 8 aaggggcggc gctggccgga ctctctcggg tggactcccg gagcggcggc ggcggcggcg 60 gtggcagact gcgagtcacg tgcgacagag gaggcggagc rcggcggccg ggtgggtgga 120 gaaaacaaag cccccagctg tcagcagagg aa 152 <210> 9 <211> 200 <212> DNA <213> Homo sapiens <400> 9 attatataaa tgaaactggt ttaggatgtg tctttttttc ccctctcttt taccttaact 60 caggttcaag cctttcttaa gctctggtaa cttgcatata yaacaaagtt gaaataaaac 120 agatttttat atagggaaag agagatgagg aagggaacat atccagtttt ctcactgtga 180 aacagagtta gcaaggaagt 200 <210> 10 <211> 201 <212> DNA <213> Homo sapiens <400> 10 gattatgtta acagaaattc tcagcctgtg tgatttgtca tattttaata tttcaagcca 60 ccattgcagc ttcttttagg tttggtttgt ctgaggcacc rcagcgagtg catgagcaca 120 cagactttga atttgtataa gtgaatggct cttccactta ctttgtggct ttgggctcat 180 cacttaactc attgatataa a 201 <210> 11 <211> 200 <212> DNA <213> Homo sapiens <400> 11 tgtgcagcaa tagtaactgg aacaacattt ctgcgatggg cagaagtggc aaggattgta 60 actcattaga ctcagaaggc gaggtcacct gagtaagcag kgtccttagt ggaacagtgg 120 gtgtaagagg aaggagagat gggtgggtgt ggggcttctt gccatctaca tatgtagggt 180 ctgcctttta cttgcacatc 200 <210> 12 <211> 166 <212> DNA <213> Homo sapiens <400> 12 gatgatactt agcttctgcg tcttcagtaa actcatctgt acaatggcct gttaaaccta 60 gctatttgaa aacatcagct aaaactaagt gatttttttg rtattcattt aggtctttta 120 tttcttgata tgcttgaaat ctaattgtct tttaaatgca ctaata 166 <210> 13 <211> 151 <212> DNA <213> Homo sapiens <400> 13 gattacaggc gtgagccacg gcacctggag agaaaaaagc ttctttaatg acgacctcaa 60 cataatttac atggagaagt ttcttaggtg gaaaaccgat ytgtatggtc ctcagaatgt 120 aacctacagg ttcttgccct cagaaaaatg c 151 <210> 14 <211> 200 <212> DNA <213> Homo sapiens <400> 14 taaggaagag ggacagctat gacctaatgc ttgcttggac cactataagc atgccaggga 60 aaatattcag gctaaattgt cggctctaag aacatgaagt rtattgattt ctttattaca 120 gctagcagat atttaagaat gttagcacag gtctttgaat aaattttgct tttaagagaa 180 gttactattt attcctaatt 200 <210> 15 <211> 200 <212> DNA <213> Homo sapiens <400> 15 tggacaatgg gaataaagca tttctaaagc accgactgga gaggaaggca acagagacaa 60 ggagagaagc cgagagacat gtctgcgtgc tgccacgcat ytgagcgatt gctctgtgaa 120 gagttgtaca ctgaacactt tcaggggagg ctgtttaccc aggcaatgtc ctcaaacaag 180 cctgtgccgg ggtgtcctgg 200 <210> 16 <211> 200 <212> DNA <213> Homo sapiens <400> 16 agaggcggcg gctccggaac gagctgctcc cacagcccct ccgctcctcg ccgtctcgcc 60 ttcgtcccca gcttacccag ttctgcgcgt tattgctcag yttgactttc ttgcacaggc 120 tcccggggat ctccatgacc gcggagcgcg ctgagcagcc ggggttctct gcgccgggan 180 ggtagcaaga ggagggcagg 200 <210> 17 <211> 200 <212> DNA <213> Homo sapiens <400> 17 ttagcagtcc cgcaaagctt gaggtttact catttttact ttctttagtc tctgggcaag 60 gaactaagag gtagtataga attctagacg atggaaactt yagaatggac aggattttaa 120 aaattatata ttccattctc ctgggttaca ggtggtctta cagctaaact atttgagact 180 gttgaagcca ttcgattttt 200 <210> 18 <211> 200 <212> DNA <213> Homo sapiens <400> 18 ttatgaggaa cagatgagat agtgaggttg agtgccagca cagtagccag tactcaatca 60 aatataactg taaaacaaag gggcagttcc taaggcccaa ratggagttt tagtcaaaag 120 gaaataaggc aaaaggccag ttacaagatg gagcaggccc tgcagtggag ttgaaaaccc 180 aggtcaccat gatgactgtg 200 <210> 19 <211> 200 <212> DNA <213> Homo sapiens <400> 19 ttaggaaatt ttcaatggag aaaataagtt ccaaaagtca ttctttctca gattaagatg 60 tcttgggttt cagtttactt ctttaaagaa agattcagag ytagtactga ggacccagac 120 tttcttgatg tgggaaatag ataattttct gttctgttga cattttttct ctctcttctc 180 tcattttcaa agtacagttc 200 <210> 20 <211> 200 <212> DNA <213> Homo sapiens <400> 20 taccatcctc acatcttgta tagactcctt ttatcagggc ctggtggtcc cagttcttcc 60 tagagaaacc catagcaaca ttcttgatgg tcagtggtcc rtggactggg ggggtcccca 120 tggccatctg tccttcctct tggctccctt ctgaggggca ggcagggtcc taaaaggaca 180 gagccctgtg gacctggcag 200 <210> 21 <211> 173 <212> DNA <213> Homo sapiens <400> 21 aaacattgtt tacataagta aacaagccag ggctgctgtg aaagttatta tggaagatac 60 aatttatata ttttctggaa aatatatcgc ctcatatata yagacaaaac ctttgcttta 120 acatgtttgt gcaactaatt acttactttc caatgagtac aatttcccac tag 173 <210> 22 <211> 200 <212> DNA <213> Homo sapiens <400> 22 gaggtgggca ctggtggctt ccagcctagg gctgtgactc tgctgttagg ggcacgctgc 60 tacacaggca ccaacctctc tagcacaggg gttcctccaa ktgtgctgtg ggggcccccg 120 agaccctttc aggggttccc ccaggttcct tgaaaggctc cnctgattag atcaggccca 180 ccaagatcac ctccctttgg 200 <210> 23 <211> 201 <212> DNA <213> Homo sapiens <400> 23 agagtcataa gaagggtttt ctctctccta aggaggaacc aaagggggaa aaaaagtctc 60 tttgcttcta gacgttgttt ctgtagatgt aaaggctgtt macttctgca gtcatcttgc 120 tgagaataaa ttcagaatga agatggcaaa atggagcaag aaataagtta atgacccgga 180 aggcctcctt ctgaagttgt t 201 <210> 24 <211> 200 <212> DNA <213> Homo sapiens <400> 24 caccatttat ttcctatttt ctctcctgag ttaaatagga aacatgtctc gcattacttg 60 aaaaatcaag tcaaactatg ctcttactag gagttatggt yctttttatg tcttagatga 120 tgcttgatct agatgaatgc ggacttgctg tagctagata aatacaatgg gagtttgaag 180 gtgtttcgta gccctggaaa 200 <210> 25 <211> 200 <212> DNA <213> Homo sapiens <400> 25 agaactgtgg agcgattcct gatttttgag caggaagagt gacaattcaa aacagtattt 60 gactagattc acggctccgt agcatcccct tgggtgggag sgggaaggct gactaggacc 120 tctgattctt ctttccctga gctttgaagg ctctgaaaat acagctgggg ggacttgccc 180 agttttctta ttaagcaatt 200 <210> 26 <211> 200 <212> DNA <213> Homo sapiens <400> 26 agcacagttc caagatgcgc tgacatgttc tgcaggcgag agagcctttc gctggactgt 60 gactcacgga ggtggtgagg ccgctggttc taaccaagag ratgccagcg agaccactca 120 gcctcccctg gtggctgccc tggtttaatg ctcatgaaat gccagcctgt ggccattctg 180 ctgggtaaag agggtggtgt 200 <210> 27 <211> 125 <212> DNA <213> Homo sapiens <400> 27 cccagcagga acccttctgg aagggaatat ttgcagagat ttcccagatc tatgtgccct 60 tctgcttcct ctatgcttac tgctctcacc tgggccaccc mtgaaggtgg gcttgctgtt 120 tcagc 125 <210> 28 <211> 200 <212> DNA <213> Homo sapiens <400> 28 tggcgcgtgt gcaattagca atgcagtgta tatacaagaa gccatgctgt taacagttta 60 tctcaagtaa tttggtgtca tttacaaaag gaagaaattc ytgccatcag aagggacaga 120 aggagatgga atgatcaaat cacagagaaa cactaaaatt actaaatcta caacagctcg 180 ccttattttt cttggaccca 200 <210> 29 <211> 200 <212> DNA <213> Homo sapiens <400> 29 gagacaacgg ggatcaggca gaggggtggg cacataggag gcaacgtaaa ccagaactga 60 ggctgggcca ggccagggtg tcagaggctg aggcggggag kcgggctgcc cttgggctgg 120 attgcaaggc gaagaagcag gagggagagg ctgagagtgt ggaggggagg gggcttgtag 180 cgtgtgattg gagggaggac 200 <210> 30 <211> 200 <212> DNA <213> Homo sapiens <400> 30 ctacccacaa aagtttcttt gtacttattt acnagtcctg atgtttttgt tagaaaaaca 60 ttgccacctn tttttaccca gccaggtctc agcctaagtg ycactgggtc cagtttctct 120 catcttctga gcagttggcc taaagtgacc ctatgttttc cttttttctt tctcctcttc 180 ctcttccaca ccagtgcaaa 200 <210> 31 <211> 200 <212> DNA <213> Homo sapiens <400> 31 tttttaggct taaaaagata ttttaatgta actctatcca attctcaatt ttaggcctgc 60 cagaattgaa actcaagctt aactataatc atcaccaagc rttacttttg aagccctccc 120 gtaaatatga ggctgttaag gatgcaatac tataccgcgt gtctcttagg ttagttggaa 180 aaatgagtta gaaaatgagg 200 <210> 32 <211> 200 <212> DNA <213> Homo sapiens <400> 32 aaagctgagt aagcacctat agtaacaggt aacaaagtat tctagggatc agctgctgtg 60 agttagttca caagtgcttt aagtacgtgt taatcaaaga ygatgtattt ttcatttctg 120 acaataaaca tctggggcat cctacatttt tcctgggcaa tttgccttta tcattatatt 180 ttaatctctg atgcaatctt 200 <210> 33 <211> 200 <212> DNA <213> Homo sapiens <400> 33 ccacttccct ctcacacaga atagatttcc atagggtgag aattcaagag aaacttttta 60 attgtacagc aaagtgttta ctttgccacc aacctgccct rtagcttaag catatggcta 120 ccttttctat caacagctgg gcttctgtag gtttggcact cggtatacca tcccagcgca 180 gtcaaaaacc tgcaggtgac 200 <210> 34 <211> 200 <212> DNA <213> Homo sapiens <400> 34 atgttgttca gagggctatt cagagaatgg aaacccaagg ctcacctttc ttgagtctaa 60 gtacccactc agccaagtat atttagcaaa atagaaaaac yaaactgctt tggtgactgg 120 actttgccaa tgtctgaaag aaagaagtaa cagaaaggag agtctaaagt tgatcctgta 180 cagtataaaa catttcccca 200 <210> 35 <211> 200 <212> DNA <213> Homo sapiens <400> 35 tggatgtact catagagagg gctggagcgc actgggacac tggcaaactg atgagtgccg 60 ggctccagaa gccgcccatg gcccttccct tcttgggatt ktatctcccc gacttggcgg 120 tgcctctgtt cctccttctg ctggcggcgg atgtgttctt cccgtaattt cctttcccgt 180 tcctggcgac gtttctgctc 200 <210> 36 <211> 200 <212> DNA <213> Homo sapiens <400> 36 gggaataaat tagttttcct tgacagtata tactgcctgt ttctgtccat cctcttattt 60 gcttcttttt ctttgctctt ccactccagt ccttaagcga ktttagtttt attcgttcct 120 cttgtccctt ttcatgataa caatgtgtca tagtctgctt taaagttttt catacttacc 180 gatgttagtt ttattgctag 200 <210> 37 <211> 200 <212> DNA <213> Homo sapiens <400> 37 tggggacaga gcctgtgggg agtcttgatg tagtcttgtc cttgaaatcc attcacggct 60 ctgctgagat ctcgggaaag agtctcttta taaatatggg kttttgtttt ttcaaatttt 120 ggaaaagtca cctctggttt taaaacaatt acttcccgct ccaaacagat tagtcacttc 180 ccaagaacca tgcttctctc 200 <210> 38 <211> 200 <212> DNA <213> Homo sapiens <400> 38 attatagaca agcagagagg gaatggatcc gagggtgacc tggctgaatt aaaagtactg 60 catctaagtg tgtgcaggtt gtagtccaca gtaatggtgc yattcggagc tgtgtgtacc 120 tgcactactg catcaggtga ccttatccag cataacggta caacagaaat ttaacttgaa 180 acaggtttat tgacttgctt 200 <210> 39 <211> 201 <212> DNA <213> Homo sapiens <400> 39 gtgacaaacc tgagactaga aaactactac atatgacatc ctacacactg actcccttct 60 tcagctctgt ctgataaagg gatgagagat cgtttcttat yattacacat acacgccact 120 tccgttcctt ccatctaaac ctagttcaga aacaccgcta agaattccca cccccaaaaa 180 aatggggatg ggggaagaga a 201 <210> 40 <211> 200 <212> DNA <213> Homo sapiens <400> 40 actgaggtta gtctcttgta ttaaactctt cacaaaatct gtttagcagc agcctttaat 60 gcatctagat tatggagctt ttttccttaa tccagctgat sagttacagc ctgttagtaa 120 catgagggga cattttggtg agaaatggga cttaactcct tccagtgtcc ttagaacatt 180 ttaattcatc ccaactgtct 200 <210> 41 <211> 200 <212> DNA <213> Homo sapiens <400> 41 cgagactcaa gataatgtac taaagacctt tagcacaatg tctgacatct ggtgttcaat 60 aagtggtgat attgctcaat ttcattccaa ttcaaactta yaaacatcca tgggaagtct 120 gctttataga caagcaatag gggagcacag taattatttg ggaggggaaa tctcagcagt 180 tcttccaagt gacacattct 200 <210> 42 <211> 200 <212> DNA <213> Homo sapiens <400> 42 atttgctgac tttgccaagc gatgagccac acaggattgg gaagcctgaa tgtcatcagt 60 caaggcccag agtgctgaac ctatgaggcc tggggagacc rctgtgaatt cctgcatcgt 120 cactgagatc atcatatcct gtgaaaaaca tctgccagcc actgtctgtg ttgtagaaag 180 ctgctgatgg gaaaacttgg 200 <210> 43 <211> 200 <212> DNA <213> Homo sapiens <400> 43 cataaaccct caataaacct aagctactat tattaagtac tgtgtgtcca cacaaaaaaa 60 aggtctggaa agatatagcc accaggagta gttatgagga rtgaggcaga gaggctgatg 120 gaggacttcc attttctatt ttgtatacta cttttctagt gtttacaaat gagcttgtat 180 tgtttttatc atcagcaaaa 200 <210> 44 <211> 200 <212> DNA <213> Homo sapiens <400> 44 atagcaggcc agggctctgt aggcttgttt tgctgctgtg cctgttgcca aatgcttgaa 60 aatcctgaag tacaaacata acagtctgaa ttaaatttca ytgattttat ctttctcttc 120 cttttctcac tgtggtgcag ttgttaatgg aattctagcc aaaaggatgt gggaggagag 180 gaaggagcca caaagggaga 200 <210> 45 <211> 200 <212> DNA <213> Homo sapiens <400> 45 ttagtcatga tgttctggtt tttatttata cattcatctg aaagagcaaa gtggtagagc 60 taaaatcagt aagcttggct ctgtttccaa actcataaaa rtgcaatatc gtggcagaaa 120 gattcttgtc ctatttctta ttttacctgg gcttttggga tttcacatgg acatagttgc 180 aaaataatac tccccaaact 200 <210> 46 <211> 200 <212> DNA <213> Homo sapiens <400> 46 ctaaccaagc ttgcccatat ttcacatgtt gagtttgttc attttaactg tcaattggcc 60 taattttccc atttctagca ttttaatttc tccagagctt kcatactaca ttgaacacca 120 attcagtgac ctggaacaat tccactcgca tttgttcctt ttagaccatt tcactgggag 180 tatgtttcca atccacttat 200 <210> 47 <211> 200 <212> DNA <213> Homo sapiens <400> 47 cagagacctg gtcctaaatc tgtgtgactt ttgcgaaatc cttttgcctc acagagctct 60 aggttcttat ctgtcaaaca ggaaccagac catcagatcc rtagagatcc aaccaatgaa 120 ggagacttcg aaaacttcac attgccgtgt caatgtgagg ttgctggatg gttatgacaa 180 gaaatcacat taaattcact 200 <210> 48 <211> 200 <212> DNA <213> Homo sapiens <400> 48 gcaggaggaa gggtgcctga ttcacagggg agatgcctga tggcgggtgg ggggctgctg 60 ttggcatccc cactggaacg tttgtcctaa tgggggcctg rctccctcat cagtggcttg 120 tcccaaaaca acaaccagtg ttcccaaacc tatacttcct ctttggattt ttgtttttca 180 tctagttgga gggaataaaa 200 <210> 49 <211> 200 <212> DNA <213> Homo sapiens <400> 49 tcattagttt attgcatccc caaggctttg ataacaaatg aagcaaatat tttcaaattt 60 ttctggttac ccaataagaa aaatcatgtg actttccaca yctgccatca ttggaagtga 120 caaaactcta gatacttggg tcattgttat catttgtcta tcttccttgg tatgctctat 180 tcttcgttgt ggagtaataa 200 <210> 50 <211> 200 <212> DNA <213> Homo sapiens <400> 50 ccagggtcgg cgcctgcccc agcctgagta ttgccctgat tctctgtacc aagtgatgca 60 gcaatgctgg gaggcagacc cagcagtgcg acccaccttc rgagtactag tgggggaggt 120 ggagcagata gtgtctgcac tgcttgggga ccattatgtg cagctgccag caacctacat 180 gaacttgggc cccagcacct 200 <210> 51 <211> 200 <212> DNA <213> Homo sapiens <400> 51 tctgacattc tctggttata aaactagagt atcaaaatgc atgagagtgc ctgctcagat 60 aaccatcaga tgaatatgaa gcttgctcat ccctgggagt rcagaaatca gctcctttta 120 cctcttgaag aagtggatac agcatcctct actgctcaat tgcataaacc agcacttgtc 180 cttggtgagc cgttgtttgc 200 <210> 52 <211> 200 <212> DNA <213> Homo sapiens <400> 52 atagaatttt tggtatactg aacaaaggga ttaaaaaggt cagttttatg tagggacttt 60 ctttgctggg gagggaaggt ctcatcccca agaattattt mtttattttc ccttcccaga 120 gtttttacat ggaagtatga aatattacct tcctactata taattaatag aaaaaataaa 180 cagagcccag attatactat 200 <210> 53 <211> 200 <212> DNA <213> Homo sapiens <400> 53 gcatagtact cagactcatt gaaagtttat agaatttggc ctgtgtggaa aactctgtgc 60 tccaagtaca acaactaact gaattctcta atttaaacaa stttgaaggg aagtgaaagg 120 ttataaaatg atacagactc ataccgcaga atcaccttaa aatgcagctg ttggagaaaa 180 aaaaaaagga agctttatgc 200 <210> 54 <211> 176 <212> DNA <213> Homo sapiens <400> 54 ttttcctggg cttcatgggc catttaaatt gtgaaccccc agggacacag tgcttagccc 60 aagccatcag tgcttgaaaa taggcatttc tgtcatcgag wttctgtatg ttgagatata 120 ttttgagata ggcttaaggg tgaggacatt tggctggagt cagcaataag ttgcaa 176 <210> 55 <211> 201 <212> DNA <213> Homo sapiens <400> 55 cacaagacta gattgcagga agatgtgaaa gcatttggct aattctgaca tttggctaat 60 aacatcctca cagtatgtcc caagtatctt ctgcccttcc yaccacggtg agcagtcatg 120 acagtgaggg tggcataggt gggatgtgtg gctggcagaa gccagtgggc aagagttgtc 180 cacagaatag tccatgagct t 201 <210> 56 <211> 200 <212> DNA <213> Homo sapiens <400> 56 tagagggggt gcgggacggg gactcacagg agatgcagga cggcccgaac atagtaattc 60 ctggtaaagg gcccgaacag cttcaccacg gcggtcatgt rcttctgtcc cctgggggag 120 ggaggaaggc gagacggcgc ggctgggcct ctcccactcg ggactccttt gctgccctgc 180 tgaccacccc agggcaccca 200 <210> 57 <211> 201 <212> DNA <213> Homo sapiens <400> 57 gttcacttgc tgctaccctc ttgtgcccac tttggcttaa taaatcccaa tccagcctag 60 ctgatttact gaagaacaaa gggatgacta gtttttgcta ygccaaggtg agtctaggat 120 cttaaaactt ttctaagggt gccaaggagt tagaatcagg ggacctaagt tttttctcct 180 gaaggcattt ctgagtagta g 201 <210> 58 <211> 201 <212> DNA <213> Homo sapiens <400> 58 tgttgatgag aaaattactt tcccttctga cattgatcct caagttttct atgaactacc 60 agaagcagta caaaaggaac tgctggcaga gtggaagaga rcaggatcag atttccacat 120 tggacataaa taagcatatt cagcaaaaag gtctgaaaag caagggaata ccattatttt 180 cggattagcg gtttattaag c 201 <210> 59 <211> 201 <212> DNA <213> Homo sapiens <400> 59 atgtgcatgt agccctttca gtaaggagtg tatggggagc ttgggccctc tgtggccttc 60 catttatttg catgtagcct cctagtcaac tagggttgta mggggagcct atgtagtccc 120 tctttggttc tctcatttcc aaatctcctg gtgacatatc tggctggtcc actgatctgt 180 ccttcacccg aaccacgact g 201 <210> 60 <211> 201 <212> DNA <213> Homo sapiens <400> 60 gcacgtgggc ctccacgatt ccttggcatg cgtgtcactc tgggcaacag gctggtcggc 60 cacgactgcg caggcgagag gcacagagcg actggagact ktagtccccc ggtttccctg 120 gagaccaggc ggaagcggcc ggaagtagcg ctgcggttgg cagcggcggg atggagaagc 180 tgggggtgga gccggaggag g 201 <210> 61 <211> 30 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 61 acgttggatg ccaagtaggg aatcaggatc 30 <210> 62 <211> 30 <212> DNA <213> Artificial Sequence <220> <223> Homo sapiens <400> 62 acgttggatg ggtctccact gatatttggg 30 <210> 63 <211> 24 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 63 ggatctttca tctcattatt aggt 24 <210> 64 <211> 30 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 64 acgttggatg gttagaaagc agcactgtgg 30 <210> 65 <211> 30 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 65 acgttggatg tcttaggagg tcattggtgc 30 <210> 66 <211> 17 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 66 tggcagctct cagaagg 17 <210> 67 <211> 31 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 67 acgttggatg gtaggattct accttagaag c 31 <210> 68 <211> 30 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 68 acgttggatg taaggagttt cagtgcaccc 30 <210> 69 <211> 17 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 69 accttagaag cttgggg 17 <210> 70 <211> 30 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 70 acgttggatg ttccagtcct agtgaaacgg 30 <210> 71 <211> 30 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 71 acgttggatg accctggtgt ctgacttttg 30 <210> 72 <211> 19 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 72 aacgggagag aaagaaggt 19 <210> 73 <211> 31 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 73 acgttggatg cacagttgta aagactttgc c 31 <210> 74 <211> 30 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 74 acgttggatg gtggtttgat aatttccaag 30 <210> 75 <211> 20 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 75 tgcctaattt ctggcaacta 20 <210> 76 <211> 31 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 76 acgttggatg gatggactta agagatcatc c 31 <210> 77 <211> 30 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 77 acgttggatg tgaacatcag cagctttccg 30 <210> 78 <211> 17 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 78 agagatcatc ccaacct 17 <210> 79 <211> 30 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 79 acgttggatg acacttgttt gatgcctgcc 30 <210> 80 <211> 30 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 80 acgttggatg tttacagact gtcctctgcg 30 <210> 81 <211> 18 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 81 ctgcctctag ggcaatga 18 <210> 82 <211> 30 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 82 acgttggatg agactgcgag tcacgtgcga 30 <210> 83 <211> 30 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 83 acgttggatg gctttgtttt ctccacccac 30 <210> 84 <211> 17 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 84 gacagaggag gcggagc 17 <210> 85 <211> 30 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 85 acgttggatg aagcctttct taagctctgg 30 <210> 86 <211> 30 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 86 acgttggatg cttcctcatc tctctttccc 30 <210> 87 <211> 21 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 87 agctctggta acttgcatat a 21 <210> 88 <211> 30 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 88 acgttggatg tttcaagcca ccattgcagc 30 <210> 89 <211> 30 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 89 acgttggatg tcaaagtctg tgtgctcatg 30 <210> 90 <211> 17 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 90 ggtttgtctg aggcacc 17 <210> 91 <211> 30 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 91 acgttggatg ctcattagac tcagaaggcg 30 <210> 92 <211> 30 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 92 acgttggatg ccatctctcc ttcctcttac 30 <210> 93 <211> 19 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 93 aggtcacctg agtaagcag 19 <210> 94 <211> 30 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 94 acgttggatg aatggcctgt taaacctagc 30 <210> 95 <211> 32 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 95 acgttggatg gcatatcaag aaataaaaga cc 32 <210> 96 <211> 23 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 96 gctaaaacta agtgattttt ttg 23 <210> 97 <211> 30 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 97 acgttggatg tttctgaggg caagaacctg 30 <210> 98 <211> 30 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 98 acgttggatg tggagaagtt tcttaggtgg 30 <210> 99 <211> 21 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 99 ttacattctg aggaccatac a 21 <210> 100 <211> 30 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 100 acgttggatg tcaggctaaa ttgtcggctc 30 <210> 101 <211> 31 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 101 acgttggatg caaagacctg tgctaacatt c 31 <210> 102 <211> 22 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 102 gtcggctcta agaacatgaa gt 22 <210> 103 <211> 30 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 103 acgttggatg cagtgtacaa ctcttcacag 30 <210> 104 <211> 30 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 104 acgttggatg acagagacaa ggagagaagc 30 <210> 105 <211> 20 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 105 cttcacagag caatcgctca 20 <210> 106 <211> 29 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 106 acgttggatg ctccgcggtc atggagatc 29 <210> 107 <211> 30 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 107 acgttggatg ttacccagtt ctgcgcgtta 30 <210> 108 <211> 17 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 108 ctgtgcaaga aagtcaa 17 <210> 109 <211> 30 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 109 acgttggatg ggcaaggaac taagaggtag 30 <210> 110 <211> 30 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 110 acgttggatg acctgtaacc caggagaatg 30 <210> 111 <211> 18 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 111 tctagacgat ggaaactt 18 <210> 112 <211> 30 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 112 acgttggatg acagtagcca gtactcaatc 30 <210> 113 <211> 30 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 113 acgttggatg tggccttttg ccttatttcc 30 <210> 114 <211> 17 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 114 cagttcctaa ggcccaa 17 <210> 115 <211> 30 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 115 acgttggatg tcaagaaagt ctgggtcctc 30 <210> 116 <211> 31 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 116 acgttggatg ctcagattaa gatgtcttgg g 31 <210> 117 <211> 17 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 117 ctgggtcctc agtacta 17 <210> 118 <211> 30 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 118 acgttggatg ctagagaaac ccatagcaac 30 <210> 119 <211> 30 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 119 acgttggatg caagaggaag gacagatggc 30 <210> 120 <211> 17 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 120 ttgatggtca gtggtcc 17 <210> 121 <211> 31 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 121 acgttggatg gtgggaaatt gtactcattg g 31 <210> 122 <211> 30 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 122 acgttggatg tctggaaaat atatcgcctc 30 <210> 123 <211> 20 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 123 ttaaagcaaa ggttttgtct 20 <210> 124 <211> 30 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 124 acgttggatg acaggcacca acctctctag 30 <210> 125 <211> 30 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 125 acgttggatg ggagcctttc aaggaacctg 30 <210> 126 <211> 20 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 126 tagcacaggg gttcctccaa 20 <210> 127 <211> 30 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 127 acgttggatg gcttctagac gttgtttctg 30 <210> 128 <211> 30 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 128 acgttggatg tgctccattt tgccatcttc 30 <210> 129 <211> 19 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 129 tgtagatgta aaggctgtt 19 <210> 130 <211> 30 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 130 acgttggatg acagcaagtc cgcattcatc 30 <210> 131 <211> 30 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 131 acgttggatg caaactatgc tcttactagg 30 <210> 132 <211> 23 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 132 agcatcatct aagacataaa aag 23 <210> 133 <211> 30 <212> DNA <213> Artificial Sequence <220> <223> primer <133> 133 acgttggatg gaatcagagg tcctagtcag 30 <210> 134 <211> 30 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 134 acgttggatg ttgactagat tcacggctcc 30 <210> 135 <211> 17 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 135 tcctagtcag ccttccc 17 <210> 136 <211> 30 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 136 acgttggatg gtgaggccgc tggttctaac 30 <210> 137 <211> 30 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 137 acgttggatg tgagcattaa accagggcag 30 <210> 138 <211> 20 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 138 ccgctggttc taaccaagag 20 <139> <211> 30 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 139 acgttggatg tctatgtgcc cttctgcttc 30 <210> 140 <211> 28 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 140 acgttggatg gctgaaacag caagccca 28 <210> 141 <211> 18 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 141 ctctcacctg ggccaccc 18 <210> 142 <211> 30 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 142 acgttggatg atctccttct gtcccttctg 30 <210> 143 <211> 30 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 143 acgttggatg gaagccatgc tgttaacagt 30 <210> 144 <211> 17 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 144 tgtcccttct gatggca 17 <210> 145 <211> 30 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 145 acgttggatg acgtaaacca gaactgaggc 30 <210> 146 <211> 30 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 146 acgttggatg ttcttcgcct tgcaatccag 30 <210> 147 <211> 20 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 147 tcagaggctg aggcggggag 20 <210> 148 <211> 30 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 148 acgttggatg tcactttagg ccaactgctc 30 <210> 149 <211> 30 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 149 acgttggatg tttttaccca gccaggtctc 30 <210> 150 <211> 17 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 150 agaaactgga cccagtg 17 <210> 151 <211> 30 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 151 acgttggatg cctcatattt acgggagggc 30 <210> 152 <211> 30 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 152 acgttggatg aggcctgcca gaattgaaac 30 <210> 153 <211> 17 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 153 gagggcttca aaagtaa 17 <210> 154 <211> 30 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 154 acgttggatg aatgtaggat gccccagatg 30 <210> 155 <211> 30 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 155 acgttggatg cagctgctgt gagttagttc 30 <210> 156 <211> 23 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 156 ttgtcagaaa tgaaaaatac atc 23 <210> 157 <211> 30 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 157 acgttggatg tgtttacttt gccaccaacc 30 <210> 158 <211> 30 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 158 acgttggatg agtgccaaac ctacagaagc 30 <210> 159 <211> 17 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 159 tgccaccaac ctgccct 17 <210> 160 <211> 30 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 160 acgttggatg cattggcaaa gtccagtcac 30 <210> 161 <211> 30 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 161 acgttggatg agtctaagta cccactcagc 30 <210> 162 <211> 17 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 162 agtcaccaaa gcagttt 17 <210> 163 <211> 30 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 163 acgttggatg atgagtgccg ggctccagaa 30 <210> 164 <211> 30 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 164 acgttggatg agaaggagga acagaggcac 30 <210> 165 <211> 17 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 165 cttcccttct tgggatt 17 <210> 166 <211> 30 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 166 acgttggatg tctttgctct tccactccag 30 <210> 167 <211> 30 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 167 acgttggatg gaaaagggac aagaggaacg 30 <210> 168 <211> 20 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 168 ccactccagt ccttaagcga 20 <210> 169 <211> 30 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 169 acgttggatg tgagatctcg ggaaagagtc 30 <210> 170 <211> 30 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 170 acgttggatg aaaccagagg tgacttttcc 30 <210> 171 <211> 24 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 171 aaagagtctc tttataaata tggg 24 <210> 172 <211> 30 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 172 acgttggatg tggataaggt cacctgatgc 30 <210> 173 <211> 30 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 173 acgttggatg agtgtgtgca ggttgtagtc 30 <210> 174 <211> 17 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 174 tacacacagc tccgaat 17 <175> 175 <211> 30 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 175 acgttggatg ggtttagatg gaaggaacgg 30 <210> 176 <211> 30 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 176 acgttggatg cagctctgtc tgataaaggg 30 <210> 177 <211> 20 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 177 aagtggcgtg tatgtgtaat 20 <210> 178 <211> 30 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 178 acgttggatg ccctcatgtt actaacaggc 30 <210> 179 <211> 30 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 179 acgttggatg gcagcagcct ttaatgcatc 30 <210> 180 <211> 20 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 180 gttactaaca ggctgtaact 20 <210> 181 <211> 30 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 181 acgttggatg gtctataaag cagacttccc 30 <210> 182 <211> 30 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 182 acgttggatg gcacaatgtc tgacatctgg 30 <210> 183 <211> 19 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 183 agacttccca tggatgttt 19 <210> 184 <211> 30 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 184 acgttggatg aatgtcatca gtcaaggccc 30 <210> 185 <211> 30 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 185 acgttggatg tgatctcagt gacgatgcag 30 <210> 186 <211> 17 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 186 tgaggcctgg ggagacc 17 <210> 187 <211> 30 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 187 acgttggatg ggaaagatat agccaccagg 30 <210> 188 <211> 30 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 188 acgttggatg aatggaagtc ctccatcagc 30 <210> 189 <211> 18 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 189 caggagtagt tatgagga 18 <210> 190 <211> 30 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 190 acgttggatg cacagtgaga aaaggaagag 30 <210> 191 <211> 30 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 191 acgttggatg tgcctgttgc caaatgcttg 30 <210> 192 <211> 22 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 192 aaggaagaga aagataaaat ca 22 <210> 193 <211> 30 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 193 acgttggatg gtaagcttgg ctctgtttcc 30 <210> 194 <211> 30 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 194 acgttggatg tgaaatccca aaagcccagg 30 <210> 195 <211> 23 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 195 gctctgtttc caaactcata aaa 23 <210> 196 <211> 31 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 196 acgttggatg gtcaattggc ctaattttcc c 31 <210> 197 <211> 30 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 197 acgttggatg gagtggaatt gttccaggtc 30 <210> 198 <211> 22 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 198 cattttaatt tctccagagc tt 22 <210> 199 <211> 30 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 199 acgttggatg caaacaggaa ccagaccatc 30 <210> 200 <211> 30 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 200 acgttggatg agcaacctca cattgacacg 30 <210> 201 <211> 17 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 201 accagaccat cagatcc 17 <210> 202 <211> 30 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 202 acgttggatg aggtttggga acactggttg 30 <210> 203 <211> 30 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 203 acgttggatg ggaacgtttg tcctaatggg 30 <210> 204 <211> 20 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 204 acaagccact gatgagggag 20 <210> 205 <211> 30 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 205 acgttggatg ttttctggtt acccaataag 30 <206> 206 <211> 31 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 206 acgttggatg caatgaccca agtatctaga g 31 <210> 207 <211> 18 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 207 atcatgtgac tttccaca 18 <210> 208 <211> 30 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 208 acgttggatg agtgcagaca ctatctgctc 30 <210> 209 <211> 30 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 209 acgttggatg aagtgatgca gcaatgctgg 30 <210> 210 <211> 20 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 210 cacctccccc actagtactc 20 <210> 211 <211> 30 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 211 acgttggatg agagtgcctg ctcagataac 30 <210> 212 <211> 30 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 212 acgttggatg tcaagaggta aaaggagctg 30 <210> 213 <211> 17 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 213 tgctcatccc tgggagt 17 <210> 214 <211> 30 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 214 acgttggatg gtagggactt tctttgctgg 30 <210> 215 <211> 30 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 215 acgttggatg tgtaaaaact ctgggaaggg 30 <210> 216 <211> 20 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 216 ctcatcccca agaattattt 20 <210> 217 <211> 30 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 217 acgttggatg ctgcggtatg agtctgtatc 30 <210> 218 <211> 30 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 218 acgttggatg ctctgtgctc caagtacaac 30 <210> 219 <211> 21 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 219 aacctttcac ttcccttcaa a 21 <210> 220 <211> 30 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 220 acgttggatg ttagcccaag ccatcagtgc 30 <210> 221 <211> 30 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 221 acgttggatg ctcaccctta agcctatctc 30 <210> 222 <211> 17 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 222 gcatttctgt catcgag 17 <210> 223 <211> 30 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 223 acgttggatg acatcctcac agtatgtccc 30 <210> 224 <211> 30 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 224 acgttggatg tcactgtcat gactgctcac 30 <210> 225 <211> 20 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 225 caagtatctt ctgcccttcc 20 <210> 226 <211> 29 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 226 acgttggatg aacagcttca ccacggcgg 29 <210> 227 <211> 30 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 227 acgttggatg aaaggagtcc cgagtgggag 30 <210> 228 <211> 20 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 228 cttcaccacg gcggtcatgt 20 <210> 229 <211> 30 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 229 acgttggatg taactccttg gcacccttag 30 <210> 230 <211> 30 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 230 acgttggatg ctgaagaaca aagggatgac 30 <210> 231 <211> 17 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 231 cctagactca ccttggc 17 <210> 232 <211> 31 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 232 acgttggatg gcttttcaga cctttttgct g 31 <210> 233 <211> 30 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 233 acgttggatg aaaaggaact gctggcagag 30 <210> 234 <211> 18 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 234 tgtggaaatc tgatcctg 18 <210> 235 <211> 30 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 235 acgttggatg tgagagaacc aaagagggac 30 <210> 236 <211> 30 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 236 acgttggatg ttgcatgtag cctcctagtc 30 <210> 237 <211> 17 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 237 gactacatag gctcccc 17 <210> 238 <211> 30 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 238 acgttggatg atgcgtgtca ctctgggcaa 30 <210> 239 <211> 29 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 239 acgttggatg tggtctccag ggaaaccgg 29 <210> 240 <211> 17 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 240 cagagcgact ggagact 17  

Claims (25)

The polynucleotide of SEQ ID NO: 57, comprising: a polynucleotide comprising 8 or more consecutive nucleotides comprising the 101 st base or a complementary polynucleotide thereof; A polynucleotide of SEQ ID NO: 58, comprising: a polynucleotide comprising 8 or more consecutive nucleotides comprising the 101 st base or a complementary polynucleotide thereof; The polynucleotide of SEQ ID NO: 59, comprising: a polynucleotide comprising 8 or more contiguous nucleotides comprising the 101 st base or a complementary polynucleotide thereof; And The polynucleotide set forth in SEQ ID NO: 60, comprising a polynucleotide comprising 8 or more contiguous nucleotides comprising the 101 st base or a complementary polynucleotide thereof. delete The polynucleotide set according to claim 1, which is used for diagnosing myocardial infarction of males and non-smokers. The polynucleotide set of claim 1, wherein the eight or more contiguous nucleotides are 8 to 70 contiguous nucleotides. delete delete The polynucleotide set of claim 1, wherein the polynucleotide set is an allele specific probe. The polynucleotide set of claim 1, wherein the polynucleotide set is an allele specific primer. delete delete delete A microarray for detecting monobasic polymorphism comprising the polynucleotide set of claim 1. A kit for detecting monobasic polymorphism comprising the polynucleotide set of claim 1, 3 or 4. a) providing a nucleic acid sample isolated from the diagnostic subject; And b) determining an allele of the polymorphic site, each 101 base of the polynucleotide set consisting of SEQ ID NOS: 57-60; and obtaining data for identifying a subject having an altered risk of developing myocardial infarction. 15. The method of claim 14, Step b) includes allele-specific probe hybridization, allele-specific amplification, sequencing, 5 'nuclease digestion. ), Molecular beacon assay, oligonucleotide ligation assay, size analysis, and single-stranded conformation polymorphism. Method carried out by the method. 15. The method of claim 14, The altered risk is increased risk. 15. The method of claim 14, And said altered risk is a reduced risk. 15. The method of claim 14, c) determining whether the allele of the polymorphic site of the polynucleotide set consisting of SEQ ID NOS: 57-60 comprises a risk allele. delete delete a) contacting a test sample comprising nucleic acid molecules with a reagent that specifically hybridizes under stringent hybridization conditions with the polynucleotide set of claim 1, 3, or 4; And b) detecting the formation of hybridized double strands. 22. The method of claim 21, Step b) includes allele-specific probe hybridization, allele-specific amplification, sequencing, 5 'nuclease digestion. ), Molecular beacon assay, oligonucleotide ligation assay, size analysis, and single-stranded conformation polymorphism. Method carried out by the method. delete delete delete