KR101560706B1 - Composition for diagnosing obesity - Google Patents

Abstract

The present invention relates to an obesity diagnostic composition comprising an agent for measuring mRNA of LOC653211 gene or a protein level encoded by the gene, an obesity diagnostic kit comprising the composition, and a method for screening an obesity treating agent. The LOC653211 gene according to the present invention shows a statistically significant correlation with obesity-related markers, and the expression of the LOC653211 gene is markedly decreased as the obesity degree is increased, which is useful for the diagnosis of obesity.

Description

Composition for diagnosing obesity [0002]

The present invention relates to an obesity diagnostic composition comprising an agent for measuring mRNA of LOC653211 gene or a protein level encoded by the gene, an obesity diagnostic kit comprising the composition, and a method for screening an obesity treating agent.

In recent years, the obesity population has been rapidly spreading due to the urbanized living environment and excessive nutrition. Obesity is not only a mere external problem but also a cause of insulin resistance, type 2 diabetes, hypertension, hyperlipidemia, arteriosclerosis, fatty liver, cirrhosis It is known to be associated with an increase in the incidence of metabolic syndrome, which is a social problem. According to the 2010 World Health Organization (WHO), over 1.6 million people worldwide are overweight and about 400 million people are obese, and about 2.6 million people die each year from obesity and overweight. In 2015, 2.3 billion people will be overweight and 0.7 billion will be obese. In Korea, the prevalence of obesity is increasing by an average of about 400,000 a year. According to the analysis of the National Health and Nutrition Examination Survey data of the Ministry of Health and Welfare and the Disease Control Headquarters, the obesity rate (BMI≥ 25 kg / m ² ) increased from 26.0% in 1998 to 30.8% (36.3% in men and 24.8% in women) in 2010, and it was higher than that of normal weight in people with hypertension, diabetes, (Hypertension 2.5 times, diabetes 2 times, hypercholesterolemia 2.3 times, and hypertriglyceridemia 2.4 times). As a result of the increase in the number of patients with metabolic syndrome such as hypertension caused by obesity and diabetes mellitus, the cost of health insurance treatment has steadily increased. As of 2010, major chronic disease treatment costs account for 31% (National Health Insurance Corporation, 2010) Not only Korea but the entire world is declaring war on obesity.

Obesity means a state of excessive accumulation of fat in the body, and it occurs not only as a cause of one cause but as a combination of genetic and environmental factors. Bouchard et al. (1990) investigated whether there is a correlation between individual differences in body weight gain and genotype. In a study of identical twins, we found that individual differences in body weight and body fat change patterns for overeating were due to genetic factors. Levine et al. (2005) suggested that non-excercise activity thermogenesis is a major factor in individual differences in weight control programs, and approximately 40-70% of human obesity-related phenotypes are genetic (Comuzzie, 1998). Therefore, it is very important to identify genetic markers that can diagnose obesity quickly and accurately.

Accordingly, the present inventors have continued to develop a gene marker capable of diagnosing obesity, and as a result, the present invention has been completed by confirming the LOC653211 gene whose expression specifically decreases in obese patients with a high BMI.

It is an object of the present invention to provide an obesity diagnostic composition comprising an agent for measuring mRNA of LOC653211 gene or a protein level encoded by the gene.

It is still another object of the present invention to provide an obesity diagnostic kit comprising the composition.

Yet another object of the present invention is to provide a method for providing information for predicting the risk of obesity, which comprises measuring the level of expression of mRNA of LOC653211 gene or protein encoded by the gene from a biological sample.

A further object of the present invention is to provide

(a) culturing cells into which the LOC653211 gene has been introduced;

(b) treating the cell of step (a) with a candidate substance for increasing or inhibiting the expression of the protein encoded by the LOC653211 gene; And

(c) comparing the expression level of the LOC653211 gene with the cells treated in the step (b) in the control, and a method for screening the therapeutic agent for obesity.

In order to solve the above problems, the present invention provides an obesity-detecting composition comprising an agent for measuring mRNA of LOC653211 gene or protein level encoded by the gene.

The present invention also provides an obesity diagnostic kit comprising the composition.

The present invention also provides a method for providing information for predicting the risk of obesity, comprising measuring the level of mRNA of the LOC653211 gene or the protein encoded by the gene from a biological sample.

In addition,

(a) culturing cells into which the LOC653211 gene has been introduced;

(b) treating the cell of step (a) with a candidate substance for increasing or inhibiting the expression of the protein encoded by the LOC653211 gene; And

(c) comparing the expression level of the LOC653211 gene with the cells treated in the step (b) in a control.

The LOC653211 gene according to the present invention shows a statistically significant correlation with obesity-related markers, and the expression of the LOC653211 gene is markedly decreased as the obesity degree is increased, which is useful for the diagnosis of obesity.

FIG. 1 shows the expression level of LOC653211 gene in obesity group when compared with normal group.

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

The present invention provides an obesity diagnostic composition comprising an agent for measuring mRNA of LOC653211 gene or a protein level encoded by the gene.

The LOC653211 gene of the present invention is characterized by being represented by the nucleotide sequence of SEQ ID NO: 1.

In the present invention, "diagnosis" means confirming a pathological condition. For the purpose of the present invention, the diagnosis is to ascertain whether or not obesity develops.

In the present invention, " agent for measuring mRNA level " includes a sense and an antisense primer complementary to the mRNA of the LOC653211 gene, or a probe.

As used herein, the term "primer" refers to a primer that is hybridized under appropriate conditions in a suitable buffer solution (for example, four different nucleoside triphosphates and a polymer such as DNA, RNA polymerase or reverse transcriptase) Refers to single stranded oligonucleotides that can serve as a starting point for DNA synthesis. The appropriate length of the primer may vary depending on the intended use, but is usually 15 to 30 nucleotides. Short primer molecules generally require a lower temperature to form a stable hybrid with the template. The primer sequence need not be completely complementary to the template, but should be sufficiently complementary to hybridize with the template. The primers of the present invention can be chemically synthesized using methods known in the art such as, for example, the phosphoramidite solid support method. Further, it can be modified by methylation, capping or the like by a known method.

The term " probe " in the present invention means a nucleic acid fragment such as RNA or DNA corresponding to a few nucleotides or hundreds of nucleotides that can specifically bind to mRNA and is labeled to confirm the presence or absence of a specific mRNA have. The probe may be prepared in the form of an oligonucleotide probe, a single stranded DNA probe, a double stranded DNA probe, or an RNA probe.

In the present invention, " agent for measuring protein level " includes an antibody specific to a protein encoded by the gene.

&Quot; Antibody " in the present invention means a specific protein molecule indicated for an antigenic site, and includes both polyclonal antibodies, monoclonal antibodies and recombinant antibodies.

The polyclonal antibody can be produced by a method well known in the art for obtaining an antibody-containing serum by injecting an antigen of the gene into an animal and collecting it from an animal. Such polyclonal antibodies can be prepared from any animal species host, such as goats, rabbits, sheep, monkeys, horses, pigs, cows, dogs, and the like. Monoclonal antibodies may also be prepared using hybridoma methods (see Kohler and Milstein (1976) European Jounal of Immunology 6: 511-519) or phage antibody libraries (Clackson et al, Nature, The antibodies of the present invention can also be prepared using two full length antibodies of the full length (SEQ ID NOs: 1, 2, 3, 4, 5, 6, Functional fragment of an antibody molecule refers to a fragment having at least an antigen binding function, and Fab, F (ab ') 2, ), F (ab ') 2, and Fv.

The LOC653211 gene according to the present invention shows a statistically significant correlation with obesity-related markers, and the expression of the LOC653211 gene is markedly decreased as the obesity degree is increased, which is useful for the diagnosis of obesity.

The present invention also provides an obesity diagnostic kit comprising the composition.

The diagnostic kit of the present invention comprises one or more other component compositions, solutions or devices suitable for the assay method. For example, the kit of the present invention can be used to carry out PCR using genomic DNA derived from a sample to be analyzed, a primer set specific for the LOC653211 gene of the present invention, a suitable amount of DNA polymerase (for example, Taq polymerase) , a dNTP mixture, a PCR buffer solution, and water. The PCR buffer may contain KCl, Tris-HCl and MgCl _2. In addition, components necessary for conducting electrophoresis to confirm whether the PCR product is amplified can be further included in the kit of the present invention.

In addition, the kit of the present invention may be a kit containing essential elements necessary for performing RT-PCR. The RT-PCR kit contains, in addition to each primer pair specific for the LOC653211 gene, a test tube or other appropriate container, reaction buffer (pH and magnesium concentrations vary), deoxynucleotides (dNTPs), Taq polymerase and reverse transcriptase , DNase, RNase inhibitor, DEPC-water, sterile water, and the like. It may also contain a primer pair specific to the gene used as a quantitative control.

In addition, the kit of the present invention may be a kit containing essential elements necessary for performing a DNA chip. The DNA chip kit may include a substrate to which a cDNA corresponding to a gene or a fragment thereof is attached as a probe, and the substrate may include cDNA corresponding to a quantitative structural gene or a fragment thereof.

In addition, the kit of the present invention may be in the form of a microarray having a substrate on which the marker gene of the present invention is immobilized.

In addition, the kit of the present invention may be a kit comprising essential elements necessary for performing ELISA. The ELISA kit comprises an antibody specific for a marker protein and comprises an agent that measures the protein level. The ELISA kit may comprise a reagent capable of detecting an antibody that has formed the antigen-antibody complex, for example, a labeled secondary antibody, chromopores, an enzyme (such as an antibody) and its substrate. In addition, antibodies specific for the quantitative control protein may be included. The antigen-antibody complex means a conjugate of a protein encoded by the LOC653211 gene and an antibody specific thereto, and the amount of the antigen-antibody complex formed can be quantitatively measured through a signal label of a detection label. Such detection labels may be selected from the group consisting of enzymes, chromophores, ligands, emitters, microparticles, redox molecules, and radioisotopes, but are not necessarily limited thereto.

The present invention also provides a method for providing information for predicting the risk of obesity, comprising measuring the level of mRNA of the LOC653211 gene or the protein encoded by the gene from a biological sample.

Biological samples in the present invention include, but are not limited to, tissues, cells, whole blood, serum, plasma, saliva, sputum, cerebrospinal fluid and urine.

In the present invention, the level of mRNA expression is determined by PCR, RT-PCR, Competitive RT-PCR, Realtime RT-PCR, RNase protection But are not limited to, methods such as RPA (RNase protection assay), Northern blotting and DNA chip.

In the present invention, the level of protein expression can be measured by an enzyme-linked immunosorbent assay (ELISA), radioimmunoassay (Radioimmunoassay) or immunohistochemistry using an antigen-antibody binding reaction using an antibody that specifically binds to a protein encoded by LOC653211 gene. (RIA), radioimmunodiffusion, Ouchterlony immunodiffusion, rocket immunoelectrophoresis, tissue immuno staining, immunoprecipitation assay, complement fixation assay, But are not limited to, methods such as fluorescence activated cell sorter (FACS) and protein chip.

Through these methods, mRNA or protein level of the LOC653211 gene compared to that of the normal group (control) can be determined by comparing the mRNA level of the LOC653211 gene or the protein level of the gene in the biological sample with the mRNA or protein expression level in the normal (control) If the level of expression is significantly reduced, it can be diagnosed as obesity, and the risk of obesity can be predicted.

In addition,

(a) culturing cells into which the LOC653211 gene has been introduced;

(b) treating the cell of step (a) with a candidate substance for increasing or inhibiting the expression of the protein encoded by the LOC653211 gene; And

(c) comparing the expression level of the LOC653211 gene with the cells treated in the step (b) in a control.

According to one embodiment of the present invention, the expression of LOC653211 gene was significantly decreased in the obese group compared to the normal group. Therefore, it is possible to screen the therapeutic agent for obesity by confirming the expression control of the LOC653211 gene.

Hereinafter, the present invention will be described more specifically based on examples. It will be apparent to those skilled in the art that the embodiments are only for describing the present invention in more detail and that the scope of the invention is not limited by these embodiments in accordance with the gist of the present invention.

Example 1. Blood Collection and Isolation of Peripheral Blood Mononuclear Cells

Blood samples were collected from Korean males divided into normal group (18.5 <BMI <23), obese OA group (25 <BMI <27) and OB group (BMI> 2 7 ) according to obesity index. Separated blood was centrifuged at 1,000 × g for 15 minutes at 4 ° C to separate plasma and stored at -70 ° C until analysis.

To separate peripheral blood mononuclear cells (PBMCs), an equal volume of DPBS was added to 8 mL of isolated blood, and concentration gradient centrifugation was performed with 6 mL of ficoll-paque (GE Healthcare, Sweden) (room temperature 3000 rpm, 30 min) . The PBMC layer was separated, and 7 mL of HBSS was added thereto, followed by centrifugation at 1500 rpm for 10 minutes at room temperature to remove the supernatant. Then, 5 mL of RBC lysis buffer (Sigma) was added and reacted at room temperature for 5 minutes. 5 mL of DPBS was added and the supernatant was removed by centrifugation at 1500 rpm for 10 minutes at room temperature. After that, 5 mL of HBSS was added and the supernatant was removed by centrifugation at 1500 rpm for 10 minutes at room temperature. The pellet obtained by centrifugation was dissolved by adding 1 mL of TRIzol (Invitrogen, Grand Island, NY) and stored at -70 ° C until the total RNA was isolated.

Example 2 Measurement of Obesity Related Markers in Blood

The following experiment was conducted using the plasma isolated in Example 1 above.

2-1. Plasma Glucose Measurement

Glucose measurement solution (Asan kit, Korea) was used to measure plasma glucose concentration. 3 μl of enzyme solution was added to 200 μL of plasma, followed by reaction at 37 ° C. for 5 minutes. Then, absorbance of the kinon-type red dye produced at 500 nm was measured (VERSAmax, Molecular Devices Co, USA). The result was compared with the glucose standard solution and quantified.

2-2. Measurement of plasma triglyceride

The concentration of triglyceride in plasma was measured using Asan kit (Korea) for the measurement of triglyceride according to the principle of color development using the enzyme method of McGowan et al. (1983). To 10 μL of plasma, 1.5 mL of enzyme solution was added and the mixture was reacted at 37 ° C. for 10 minutes to decompose neutral lipids in plasma into glycerol and fatty acids by lipoprotein lipase (LPL). Glycerol formed L-α-glycerophosphate by the action of ATP and GK (glycerol kinase), which reacted with O ₂ and GPO (glycerophosphooxidase) to generate H ₂ O ₂ . After peroxidase and 4-amino-antipyrine treatment were performed to develop red color, absorbance was measured at 550 nm. The result was compared with glycerol standard solution (300 mg / dL) and quantified.

2-3. Plasma total cholesterol (TC) measurement

Plasma total cholesterol concentration was measured using a total cholesterol measurement solution (Asan kit, Korea) using the enzyme method of Allain et al. (1974). To 10 μL of plasma, 1.5 mL of enzyme solution was added and incubated at 37 ° C for 5 minutes. Cholesterol ester was converted to fatty acid and free cholesterol by cholesterol esterase to quantify both plasma cholesterol present in two forms, cholesteryl ester (CE) and free cholesterol (FC). The converted free cholesterol was converted to H ₂ O ₂ and δ 4 -cholestenone by reaction with cholesterol oxidase, and this product and H ₂ O ₂ , which is a substrate, were reacted with peroxidase, phenol and 4-amino-antipyrine to produce red color After obtaining the material, absorbance was measured at 500 nm. The results were compared with cholesterol standard (300 mg / dL).

2-4. Plasma HDL-cholesterol (HDL-C) measurement

The concentration of plasma HDL-cholesterol was measured using a HDL-C measurement solution (Asan kit, Korea). 100 μL of plasma was treated with 100 μL of the separation solution, and LDL (low density lipoprotein) and VLDL (very low density lipoprotein) containing apo B, among lipoproteins, were precipitated by the action of the tungstate and magnesium cations contained in the separation solution Warnick et al., 1982). After centrifugation, the supernatant was used to measure the absorbance at 500 nm after the color reaction by the same method as the measurement of total cholesterol concentration in 2-3 above. The results were compared with the cholesterol standard curve (50 mg / dL) to quantify HDL-C.

2-5. Plasma LDL-cholesterol (LDL-C) measurement

The concentration of plasma LDL-cholesterol was calculated according to the following formula (Friedewald et al., 1972).

[LDL-C] = [Total-cholesterol] - [HDL-C] - 0.2 x [Triglyceride]

2-6. Plasma free fatty acid (FFA) measurement

The concentration of plasma free fatty acids was measured using a non-esterified fatty acid (NEFA kit, Wako, Osaka, Japan) according to the principle of color development using the enzyme method. 1 mL of NEFA-1B was added to 25 μL of plasma and incubated at 37 ° C for 10 minutes. After that, 500 μL of NEFA-2B reagent was added, reacted at 37 ° C for 10 minutes, and absorbance was measured at 555 nm. The results were compared with free fatty acid standard solutions.

2-7. Plasma phospholipid (PL) measurement

Plasma phospholipid concentrations were measured using a kit for measuring phospholipid (Shinanyang Chemical Co., Ltd., Japan). To 100 μL of plasma, 1 mL of PL-M-1 was added and reacted at 37 ° C for 5 minutes. Thereafter, 500 μL of PL-M-2 was added and reacted at 37 ° C for 5 minutes, and the absorbance of the purple color was measured at 600 nm. The results were compared with phospholipid standard solutions and quantified.

2-8. Measurement of plasma apolipoprotein (ApoA-I, ApoB)

Plasma apolipoprotein concentrations were determined by measuring the concentration of apo A-Ⅰ and apo B using an enzyme-linked immunosorbent assay ELISA kit (ALERCHEK, INC, USA).

2-9. Results analysis

The results of Examples 2-1 to 2-8 are shown in Table 1 below.

N
(Male, n = 6) OA
(Male, n = 10) OB
(Male, n = 7) Age (year) 34.57 + 5.04 39.60 + - 4.54 33.86 + - 4.71 BMI (kg / m ² ) 21.59 ± 0.48 ^c 25.75 ± 0.18 ^b 28.47 ± 0.51 ^a Glucose (mmol / L) 4.87 ± 0.26 ^b 5.36 ± 0.16 ^ab 5.62 ± 0.24 ^a TG (mmol / L) 2.32 ± 0.09 ^b 4.42 ± 0.68 ^a 5.58 +/- 0.85 ^a Total-C (mmol / L) 4.09 ± 0.28 ^b 4.83 ± 0.19 ^b 6.24 + 0.40 ^a PL (mg / dL) 1.04 0.08 ^c 1.29 + 0.07 ^b 1.50 0.04 ^a LDL-C (mmol / L) 2.38 ± 0.30 ^b 2.99 ± 0.15 ^b 3.82 ± 0.36 ^a ApoB (mg / dL) 46.17 ± 3.80 ^b 61.24 + - 6.43 ^b 93.57 ± 6.49 ^a HDL-C (mmol / L) 0.58 ± 0.08 ^a 0.41 + - 0.03 ^b 0.40 ± 0.05 ^b ApoA-I (mg / dL) 160.49 + 13.61 ^a 139.32 ± 2.41 ^ab 134.01 + - 8.45 ^b

(* Data are mean ± SE ^abc Means are significantly different among groups at p <0.05)

As shown in Table 1, the BMI, glucose, triglyceride, total cholesterol, phospholipid, LDL-cholesterol and apolipoprotein B were significantly increased with increasing obesity, while HDL-cholesterol and apolipoprotein AI had an obesity index The higher the value, the lower the significance.

Example 3. Analysis of obesity-related genes

3-1. Isolation of total RNA

The PBMC isolated in Example 1 was centrifuged at 3,250 x g at 4 DEG C for 10 minutes. The supernatant was taken, and 200 μL of chloroform was added thereto, followed by centrifugation at 3,250 × g, 4 ° C. for 20 minutes. The supernatant was again taken, and an equal amount of isopropanol was added thereto. Thereafter, the RNA pellet was obtained by centrifugation at 3,000 x g at 4 DEG C for 10 minutes. The RNA pellet was washed twice with 75% ethanol, dried and dissolved in DEPC (diethyl pyrocarbonate) treated water. And stored at -70 ° C until analysis.

3-2. Microarray experiment

3-2-1. RNA QC and total cRNA synthesis

RNA synthesis and purification were performed using the Illumina Total Prep RNA Amplification Kit (Ambion, Austin, Tex.). More specifically, first strand cDNA synthesis was obtained by reverse transcription using 500 ng of the T7 oligo (dT) primer to the RNA obtained in the above Example 3-1, and the second The second strand cDNA synthesis was generated by converting single-strand cDNA to double-strand cDNA by reaction of DNA polymerase with RNase H. After in vitro transcription cRNA synthesis step, biotin-NTP labeled cRNA was obtained and quantified with ND-1000 spectrophotometer (NanoDrop, Wilmington, DE). In addition, the quality of the RNA was determined using Agilent's Bioanalyzer 2100 and then used in the experiments.

3-2-2. Whole gene expression microarray experiment

Microarray experiments were performed using Whole-genome gene expression with IntelliHyb Seal System (Illumina, Inc., San Diego, USA) according to manufacturer's recommendations. 750 ng of the labeled cRNA sample obtained in Example 3-2-1 was hybridized to Human HT-12 v4.0 Expression Beadchip and reacted in a 58 ° C thermostat for 16-18 hours. Detection of array signals was performed using Amersham fluorolink streptavidin-Cy3 (GE Healthcare Bio-Sciences, Little Chalfont, UK) according to the bead array manual.

3-2-3. Derive scan and extract image data

Bead chip scanning was performed by the manufacturer using the Illumina BeadArray Reader Confocal Scanner, and the bead chip image derived from the scanner was analyzed with raw data using the Gene Expression Module using Illumina BeadStudio Software (Illumina BeadStudio v3.1.3 (Gene Expression Module v3.3.8)).

3-2-4. Standardization of Microarrays

The Illumina Mouse WG-6 V2 Expression Bead Chip has been normalized for analysis. The standardization method is based on the assumption that the distribution of genes is almost the same in all time-series because the number of genes is sufficiently large. For convenience of analysis, a non-parametric method using the distribution of genes as an index of standardization Quantile Normalization was used.

3-2-5. Extraction of Differentially Expressed Genes (DEG)

BMI and 8 markers related to obesity as confirmed in Example 2 above were selected by linear regression model (P <0.05). The results are shown in Table 2 and Fig.

Phenotypei = β0 + β1 Expressioni + εi, εi ~ N (0, σ2)

Gene
symbol Negative
correlation
(-) Positive correlation
(+) BMI Glucose TG TC PL ApoA-I LDL HDL ApoB LOC653211 0.0001 0.0286 0.0032 0.0027 0.0002 0.0267 0.0409 0.0002 0.0138

As shown in Table 2, the LOC653211 gene was found to be significantly associated with obesity-related markers (p-value <0.05).

In addition, as shown in FIG. 1, the LOC653211 gene was found to have a lower expression level as the degree of obesity increased, and thus it was confirmed that the LOC653211 gene showed a negative correlation with obesity.

Through the above experimental results, it was confirmed that obesity can be diagnosed by measuring the level of mRNA of LOC653211 gene or the protein encoded by the gene, and that the substance for controlling obesity can be screened by identifying the substance that regulates the expression of the gene.

<110> Kyungpook National University Industry-Academic Cooperation Foundation <120> Composition for diagnosing obesity <130> 179 <160> 1 <170> Kopatentin 2.0 <210> 1 <211> 3346 <212> DNA <213> Homo sapiens <400> 1 accaaaaagt ccaaccatat gtaaaatggc atcacactgt aaagctcata ttacagtatc 60 aggacaattg atactttcaa cagcagcaat ggctgccagg cagcggtcct gaattgaaga 120 cttgtaaagg ggagaaaata cacaacacag aaaaagaaac ctaataaaat ggggaaacgg 180 attctctggg aggagcgtat gcaactccca aggcaaaatt cagaaagatg ttgtttcccc 240 tcaaaacaaa gagccttaat gcccagaatg ccacagttct gaatcgtgag tacctgcaaa 300 tagacagaag gaaaacggag gggatatgtg ggtgagagat gacctggatt ctcacgagaa 360 gaagggcagc caccttagag gtgcacggaa tacagtttca ggcctggtgc aactcaaggg 420 gcagttcacc attttccagt aagcccgaaa ttgacgaata aaagggactt aaagtattag 480 agagcagctc tttggaaacc cacggaaaac aaacattggg gattcaaagg atttgggagg 540 aaaggaggtg gaaatgagtc aaaagaagcc ttctcgagga taaaatcatg tgaccctcag 600 gaagagaatg cccaaggagc taagaaaagg gaggacaggt cctggggaaa agccctccac 660 tccaacaagc gggaagtgca taccaatggg tgcggggggg cccccgaagg taccgctttc 720 ggagcacccg aggcctccag ccggatgtca ggccacgcac caacgaggga ccaccgtgcg 780 gggacagtgt ccgcgcgcgg nnnnnnnnnn nnnnnnnnnn nnnnnnnnnn nnngtgggcc 840 ggaagggggg gcagggcggg ccgggagggc gccccacgga acacttacgc ggccgctttt 900 tcctcaggct ttcgacgcgc tggcgggaga ccgaggtggg gccgctgtaa gagcccagtc 960 gatcccgcct cgtgccctca gccgaggccc cgaccgcggg caccgacgcg ctgctcgccc 1020 ggcgccgcag tggctcctgc cgccgccctt cggggtcccc ggccgcctgc tccccagccg 1080 acgagacgga ggtggcggaa gccctgccgt ggcgctccat gggcgcgccg agccggcccc 1140 ctcccatccg cagcagcagt cgctgccgcc cgagcaagcc ccccagcgtc ggattccttc 1200 tcgcctcctc cgcggcagtc cagccagagc agaggcgcgc cctcggccgg ccgcctcttc 1260 cgttgcgcca acgcgcagga agccaatccg acgaggctgg ggacggccca aaggcctcgc 1320 gtccagtcgg ctccctaact cgaggaggat tgaccgtgcc tccgtccaat gagaggctat 1380 cgtgacgggg cggggagagg gtggtggagg aggggatccg ggttccgcgg aggaagagaa 1440 ggcggtgtag cccagaagag agtgatggcg gcgccgccca atggacgcgt gggtggaagg 1500 gcccgagcct cctcatccac cccctccttc cgccccccac ttgtgcatgc ctctgccccc 1560 ccgggccgcg gaactctcct ggggcgaccc cgagtcccac cccgctgctc cgtgccgtcc 1620 tcgcagctcg agccctcctc cagcccagca gtgctcccgc agcgtctggc cgctggtcgt 1680 cggcagacga aagggcttga accgcggctg accttagcct ttgtccctcg gcctagttcc 1740 gcagctgccg tcgaggccgc cagcccctcc cccagcatgt ttgggctaag ccgcctcctt 1800 tccgcctgcg ttggagagag aggtaccaga aggaatgaaa ggactggggc ttgtctccat 1860 ggcaactgaa agaaagaggt gggttataaa tccttcgcca ccctcctgtg gcgaaggagg 1920 ggaagtggga gccgggaagt agagttggcc ttttgcagga gagtctggag cccctttctg 1980 ggtgatgcat tgggggccat agaaggattt ggctgccgct ttggagcatg gatttttttt 2040 ttaatcatct tcctttctct tttggagaga aaaattggtc acctccagga ctgagaccct 2100 gcccttcca agcatcccca ctaccaaagg ctgcccaagc ccggaaggat cccatcccgt 2160 tctgtactct gaggtcactt cctcactccg tcaaatttca ctggactgtg ggctaattaa 2220 aagtgcctcc gtattagcgg ggcgtggtgg cccatgccta taatcccagc tacttgggaa 2280 gctgaggcag gaaaattgct tgaacccggg tggcggaggc tgcagtgacc caacattgtg 2340 caacaagagc gaaactccgt ctcaaaaaaa ataaaaaata aaaaagagtg cctccctggg 2400 gagactgtaa tgcccactgg gcagtgatcc taatgcccct tgggcagtga tccttattcc 2460 cctctcccat aatgattggt tgtagcaggc tgaacaaaaa ggttgagctt cgcttgtgct 2520 tgctgaatag aattctcacg cccactccac gttagctccc ctcactgtag gactagctct 2580 gaaaattcta gctcctgtag gaagccaaca cagacggatc tacaagaacc aaaaacagct 2640 atagctactc ttctaaatct cagtgcctga cattgcctct ctttctgcct cctctctcca 2700 ccccccaaaa aatgtggtcc tccttgacct gagggagctt ccgtccaagt ctgagctctg 2760 atcatcctga agaatcaaag tatcatcatc ccttggctgg caagatgaac tccaacatca 2820 aaacacttga aacagtagta tttttacaca atttgcccac tgcctctaca tctcacttag 2880 tccttgcagt aacatttgga tgatgaggta acttatccag aagcacaaag cctaaagaat 2940 gacagagctt ggactcctga gaagttcagc tactgcccgc cttatgataa ccaaaagaaa 3000 gcggccgggc gcagtggctc aactcctgta atcccagcac tttgggaggc cgaggcgtgc 3060 gaatcacaag gtcaggagtc cgagaccagc ctggccaaca cagggaaacc ccgtctctac 3120 taaaaaaaac aaacacaaaa attagttggg cgtggtggtg ggcacctgta atcccagcta 3180 cttgggaggc tgaggcagga gagtcacttg aatccggtag gcggaggttg cagtgagccg 3240 agatcgtgcc actgcactcc agcctgggag acagtgagac tccgtctcta aaaaaaaatg 3300 aataaataaa taagaaggca caccacaagt tcgagaccac ttctac 3346

Claims (10)

1. A composition for diagnosing obesity comprising an mRNA of LOC653211 gene represented by the nucleotide sequence of SEQ ID NO: 1 or an agent for measuring the level of protein encoded by the gene.
delete The composition for diagnosing obesity according to claim 1, wherein the agent for measuring the mRNA level is a sense and antisense primer complementary to the mRNA of the gene, or a probe.
The composition for diagnosing obesity according to claim 1, wherein the agent for measuring the protein level is an antibody specific to a protein encoded by the gene.
An obesity diagnostic kit comprising the composition of claim 1.
(a) measuring the level of expression of the mRNA of the LOC653211 gene represented by the nucleotide sequence of SEQ ID NO: 1 or the protein encoded by the gene from the biological sample; And
(b) determining that the risk of obesity is high when the level of mRNA or protein expression of the LOC653211 gene is lower than that of the normal group.
The method according to claim 6, wherein the biological sample is at least one selected from the group consisting of tissue, cells, whole blood, serum, plasma, saliva, sputum, cerebrospinal fluid and urine.
7. The method of claim 6, wherein the mRNA expression level is selected from the group consisting of Polymerase Chain Reaction (PCR), Reverse Transcription Polymerase Reaction (RT-PCR), Competitive RT-PCR, Realtime RT- ), RNase protection assay (RPA; Northern blotting), and a method using a DNA chip. 2. The method according to claim 1, Information providing method.
7. The method according to claim 6, wherein the protein expression level is determined by Western blotting, enzyme linked immunosorbent assay (ELISA), radioimmunoassay (ELISA), or immunohistochemistry using an antibody-antibody binding reaction using an antibody specifically binding to a protein encoded by LOC653211 gene. Immunoassay, radioimmunoassay (RIA), radioimmunodiffusion, Ouchterlony immunodiffusion, rocket immunoelectrophoresis, tissue immuno staining, immunoprecipitation assays, complement fixation Wherein the measurement is carried out through at least one method selected from the group consisting of a method using a fluorescence activated cell sorter (FACS) and a protein chip. Way.
(a) culturing cells into which the LOC653211 gene represented by the nucleotide sequence of SEQ ID NO: 1 has been introduced;
(b) treating the cell of step (a) with a candidate substance for increasing or inhibiting the expression of the protein encoded by the LOC653211 gene; And
(c) comparing the expression level of the LOC653211 gene in the control and the cells treated in the step (b).

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