한우 마블링 예측을 위한 폴리뉴클레오티드 및 그 진단방법
The invention relates to a device and method for cattle e [pul[pul] ring prediction for polynucleotide, can be used for immobilizing polynucleotide microarray gene derived ADIPOQ morphology 1 provided in detecting defects in cattle for predicting e [pul[pul] ring 258 can be polynucleotide or sequence number 1 for inserting or represented by the second base is T or C ADIPOQ gene derived for immobilizing polynucleotide capable of detecting a single nucleotide polymorphism in 73 the polynucleotide for predicting e [pul[pul] ring cattle, composition, kit and method for predicting e [pul[pul] ringe [pul[pul] ring cattle predicting polynucleotide encoded by the cattle are disclosed. The phenotype of traditional e [pul[pul] ring bred by the same measuring technique upon which the meat and the measurement value, candidate bell bull moieties through growth and conductor to assay transformed progeny, selecting candidate bell bull again looked like. However, as well as the meat is added measurements may be made using the same microsatellite, since accuracy of the evaluation based on parent-commerce of breeding was assessed by low disclosed. Also, because the many candidate bell bull breeding via two generations, reciprocating, non-breeding, facility management is performed breeding person like the signal is to render the disclosed. In order to solve such problems, cookies that can estimate the quality of the studies of genetic markers extended genetic capability reduces disclosed. E.g., PCR using the same RAPD (Random Amplified Polymorphic DNA), SSCP (Single Strand Conformational Polymorphism), AFLP (Amplified Fragment Length Polymorphism) and DNA analysis techniques such as RFLP (Restriction Fragment Length Polymorphism) livestock major economic response from the molecular marker related to screening. This molecule labels molecule markers most widely used single base polymorphisms, i.e. SNP (Single nucleotide polymorphism) sign disclosed. SNP if the SNP is transformed phenotype is affecting major economic livestock animals at a different level of transformation can be utilized as a predictive marker. The use of a marker 10 - 2012 - 0011728 SNP presents the public Internet route call (2012. 02. 08.) as, microsatellite markers useful for selecting an early indication of meat and meat amount or nucleotide sequence are disclosed. In white adipose tissue of several mammalian species (ADIPOQ) adiponectin secreted by proteins and fgfr5, etc. the adipose tissue mass and in relationship of being inversely proportionaloh D gun neck [thin[thin] secretion. Recent study, even generated oh D gun neck [thin[thin] brown adipose tissue and skeletal muscle, glucose and emulsified oil biosynthesis that neoplastic synthesis has been found. Also, fatty acid oxidation of arrayed microchambers tube tongueoh D gun neck [thin[thin] increased and glucose absorptive as insulin sensitivity regulation has been found. Such studies result in modulation of fat and a carbohydrate metabolism oh D gun neck [thin[thin] this exhibits that plays an important role. In such a technical background, the present inventors believe that microsatellite e [pul[pul] ring transformation has been developed for predicting polynucleotides and method of diagnosing. Finally, the aim of the invention are practical and enabling rapid e [pul[pul] ring prediction for predicting e [pul[pul] ring cattle polynucleotide sequences 30 to 60 seconds. It is another object of the present invention for predicting composition e [pul[pul] ring cattle are disclosed. The another object is to provide a kit for predicting e [pul[pul] ring cattle are disclosed. The another object is to provide a prediction method for predicting e [pul[pul] ring cattle e [pul[pul] ring polynucleotide encoded by bred by the same are disclosed. The advantage of and another object of the invention to form a description, claims and drawings by more clearly with each other. In order to solve such the swing, In one aspect of the present invention, sequence number 1 can be used for immobilizing polynucleotide can be inserted or in detecting defects in 258 ADIPOQ gene derived from microarray for predicting e [pul[pul] ring cattle can be polynucleotide is provided. In another aspect, the sequence numbers 1 gene can be used for immobilizing polynucleotide derived from a second base is T or C ADIPOQ 73 can be a mono-nucleotide polymorphism detection for predicting e [pul[pul] ring cattle can be polynucleotide is provided. In another aspect of the present invention, compositions are provided that the polynucleotide comprising cattle e [pul[pul] ring prediction can be. In one embodiment of the present invention, a primer set 2 and 3 characterized in that the polynucleotide sequence number. In another aspect of the present invention, the composition can be a kit for predicting e [pul[pul] ring cattle. In another aspect of the present invention, (a) extracting DNA genome of cattle; (B) the step (a) extracted from the genome DNA as templates, the sequence numbers 2 and 3 an amplifying genomic DNA for primer set mold step; and (C) amplifying the consequences to, sequence number 1 258 of second base sequence represented by sequence number 4 is not performed relative to plate is inserted polynucleotides occurs e [pul[pul] ring cattle good prediction method is provided determining rating e [pul[pul] ring cattle can be. In another aspect of the present invention, (a) extracting DNA genome of cattle; (B) the step (a) extracted from the genome DNA as templates, the sequence numbers 2 and 3 an amplifying genomic DNA for primer set mold step; and (C) amplifying the consequences to, sequence number 1 73 of second nucleotide sequence allele type is more excellent than when CT or TT CC e [pul[pul] ring grade cattle when determining e [pul[pul] ring cattle can be prediction method is provided. In another aspect of the present invention, (a) extracting DNA genome of cattle; (B) the step (a) extracted from the genome DNA as templates, the sequence numbers 2 and 3 an amplifying genomic DNA for primer set mold step; and (C) amplifying the consequences to, sequence number 1 258 of second base sequence represented by polynucleotides is inserted sequence number 4 has taken place, the sequence numbers 1 73 when a second nucleotide sequence of CT-allele, the sequence numbers 1 258 of second base sequence represented by sequence number 4 does not occur and inserted polynucleotides, when the second nucleotide sequence allele-CC or TT 73 further excellent rating determining e [pul[pul] ringe [pul[pul] ring cattle than cattle can be prediction method is provided. The ness microsatellite e [pul[pul] ring polynucleotide can be effective and practical method of predicting, existing techniques and input phenotype measurement process from a decryption key. Also, based on the same process from selecting bell right producing cattle meat quality. Figure 1 SNPs ADIPOQ gene by in one example of the present invention, insertion and defective nucleotide sequence is shown agarose gel and exhibits primer set. Below, the present invention more specifically described as follows. In one aspect of the present invention, sequence number 1 can be used for immobilizing polynucleotide can be inserted or in detecting defects in 258 ADIPOQ gene derived from microarray for predicting e [pul[pul] ring cattle can be polynucleotide is provided. In another aspect, the sequence numbers 1 gene can be used for immobilizing polynucleotide derived from a second base is T or C ADIPOQ 73 can be a mono-nucleotide polymorphism detection for predicting e [pul[pul] ring cattle can be polynucleotide is provided. The according to the invention, "polynucleotide" concern units by means leading into a nucleotide polymer which is covalently bonded chain shape, may comprise an be analogs thereof. The according to the invention, "ADIPOQ gene" includes a glucose level and fatty acid regulating the degradation that are known to adiponectin (adiponectin) gene encoding big. The disclosure to movable through the ADIPOQ gene 1, length about 11,429,556 DNA base sequences. The ADIPOQ gene nucleotide sequence database of NCBI Blast known such as can be obtained, preferably comprising a polynucleotide of seq ID no 1 can. The according to the invention, "sequence number 1" areas (polymorphic sequence) comprising the polymorphic sequences are disclosed. SNP (single nucleotide polymorphism) is obtained during polymorphic sequences comprising polynucleotide sequence comprising a sequence by big areas (polymorphic site). The poly the [thyu[thyu] the [ley[ley] five mote [tu[tu] which will grow sequences implementation being RNA or DNA. The according to the invention, the nucleotide sequence of the DNA base sequence is one "single nucleotide polymorphism" (A, T, G, C) show differences in genetic change or a mutation by big. In another aspect of the present invention, compositions are provided that the polynucleotide comprising cattle e [pul[pul] ring prediction can be. In one embodiment of the present invention, a primer set 2 and 3 characterized in that the polynucleotide sequence number. The term "primer" are used in the invention is a nucleic acid strand complementary to single-stranded nucleotide sequence and be copy means, acts as a starting point for the synthesis of primer extension products can be. The primer length and sequence is permitted to extend product synthesis of accomplishing. The primer is preferably 15 - 30 base pairs after are made the length the nanometer range. In another aspect of the present invention, the composition can be a kit for predicting e [pul[pul] ring cattle. In the present invention, if applied to a performs amplification PCR, "kit" optionally, PCR amplification reagents required, for example, buffer, DNA polymerase (e.g., Thermus aquaticus (Taq), Thermus thermophilus (Tth), Thermus filiformis, Thermisflavus, Thermococcus literalis or Pyrococcus furiosus DNA polymerase (Pfu) obtained from thermal stability), a DNA polymerase can be cofactors and dNTPs. The kit packaging or manufacturing the agent component can be a plurality of separate com part with distant [thu[thu]. In another aspect of the present invention, (a) extracting DNA genome of cattle; (B) the step (a) extracted from the genome DNA as templates, the sequence numbers 2 and 3 an amplifying genomic DNA for primer set mold step; and (C) amplifying the consequences to, sequence number 1 258 of second base sequence represented by sequence number 4 is not performed relative to plate is inserted polynucleotides occurs e [pul[pul] ring cattle good prediction method is provided determining rating e [pul[pul] ring cattle can be. In another aspect of the present invention, (a) extracting DNA genome of cattle; (B) the step (a) extracted from the genome DNA as templates, the sequence numbers 2 and 3 an amplifying genomic DNA for primer set mold step; and (C) amplifying the consequences to, sequence number 1 73 of second nucleotide sequence allele type is more excellent than when CT or TT CC e [pul[pul] ring grade cattle when determining e [pul[pul] ring cattle can be prediction method is provided. In another aspect of the present invention, (a) extracting DNA genome of cattle; (B) the step (a) extracted from the genome DNA as templates, the sequence numbers 2 and 3 an amplifying genomic DNA for primer set mold step; and (C) amplifying the consequences to, sequence number 1 258 of second base sequence represented by polynucleotides is inserted sequence number 4 has taken place, the sequence numbers 1 73 when a second nucleotide sequence of CT-allele, the sequence numbers 1 258 of second base sequence represented by sequence number 4 does not occur and inserted polynucleotides, when the second nucleotide sequence allele-CC or TT 73 further excellent rating determining e [pul[pul] ringe [pul[pul] ring cattle than cattle can be prediction method is provided. The cattle the genome DNA of muscle, skin, blood, bone, such as organ from a variety of sources and factor is shared, most preferably can be obtained from muscle or blood. The second extraction is known in the genome DNA of the present invention contrast to which, to this end commercial kit can be used but, limited to are not correct. (B) the amplification step comprises a polymerase chain reaction (PCR; comprising Realtime PCR), methylamine (ligase chain reaction) has also the advantage chain reaction, nucleic acid sequence based amplification (nucleic acid sequence a-based amplification), transcription based amplification system (transcription-a basedamplification system), strand amplification (strand displacement amplification) or Q replicase (replicase) amplifying or this invention is in the field through any other suitable method for amplifying nucleic acid molecules known the pin is. The PCR primer sequences capable of specifically binding to the target cell through the use polymerase gene target sequence from amplifier and, the most preferred gene amplification method are disclosed. (C) analyzing the genotype is allele specific probe hybridization method (allele non-specific hybridization), fluorescent in situ hybridization (FISH), molecular beacon (molecular beacons), SNP microarray (SNP microarray), limiting enzyme fragment for a long time polymorphism (Restriction fragment length polymorphism), method using the same PCR (PCR-a based methods), Flap endonuclease (Flap endonuclease), primer extension method (Primer extension), 5 '- nuclease (5' - nuclease), oligonucleotide binding assay (Oligonucleotide Ligation Assay), size (size analysis) assays, single - strand [khen[khen] It ladles, the May [syen[syen] analysis (SSCA), modified gradient gel electrophoresis (DGGE) and single-stranded it conceives leftthe characteristic law (single-a stranded conformation polymorphism) method selected from the group consisting of polymorphic but performed by, limited to are not correct. The genotype in the molecule result difference of binding sequences of single - strand base change, mobility causing different from that which, no SSCA detected by a band. DGGE analysis using a modified gradient gels, representing sequences detected by a wild-type sequence different mobility. Other techniques of the present invention relate generally to a probe or primer complementary to a nucleotide sequence comprising a SNP is measured for plural times. More detailed embodiment is less than the broadcast receiver which utilizes the present invention. But, these embodiments only exemplify a provided for, these limited by the scope of this invention embodiment will not interpreted. Embodiments 1: measuring transgenic bovine conductor In total in 1954 of cattle is inadequate and randomly agricultural associationcentral committee telecopy machines improved cattle country from embryo, country database stores (KAPE) of 9 samples are also of meat bred by the same evaluation circle randomly for special packaging of goods from Korean animal facility. KAPE material collected from conductor weight, lampwick cross-sectional area, thickness and e [pul[pul] ringetc. region includes ratings. After time CA 24, about 5g 6 been collected from around the second longest near rib breast muscle sample. Embodiments 2: genome DNA Extraction Of genomic DNA extract, muscle sample about 1g are used. After slightly compacted samples, it is possible placing pieces and extraction buffer tube, commercial kits for genomic DNA (Wizard DNA extraction kit, Promega) according to the manufacturer's guidelines of been derived. Embodiments 3: amplification (A GenBank accession number JQ775868) small primer based on the sequence of DNAstar versio 6. 0 of DNA screening program has been designed. All bovine type having sufficient in the primer set for polymorphisms screening, alignment (alignment) of reference sequences from GenBank with the support number 6 (DQ156119, EU296533, EU313339, EU492456, EU492457 and JQ775868) running of 8 with potential SNP (g. 81965420G>C, g. 81965993G>C, g. 81966286C>A, g. 81966690A>G, g. 81966775T>C, g. 81966798G>A, g. 81966940A>G and g. 81967160T>C) has been found. The present invention routes is sufficient for cattle and have variable that appears SNP (g. 81966377T>C) screening, the SNP genome comprising a portion of the primer was to design. Each forward and reverse primer (sequence number 2, nucleotide position 81966163 - 81966184) (sequence number 3, 81966410 - 81966429) on GCAGC TCTAC TTGGC ATCC CTTGA ATCAG TCGTC CTTAC CC are disclosed. 2 μl total 20 μl of volume of 10X reaction buffer (10 mm Tris, pH 8. 3, 50 mm KCl, 0. 1% Triton X-a 100, 1. 5 mm MgCl2 ), 2. 5 mm dNTO, 10pmol of each primer, 1 unit of Taq DNA polymerase (Gibco BRL, Grand Island, NY) and 50 ng genomic DNA are used. 2 minutes 95 °C after application of heat, modified for 94 °C/1 minutes, heating cooling 59 °C/1 minutes, 72 °C/1 min cycle as for polymerization, total 35 cycle has been performed (MJ Research, PT-a 200, Watertown, MA). DNA band is detected by means of a UV light has been dyed with ethynyl [...] bromide. Embodiments 4: detection of single base polymorphisms DNA samples are purified by using PCR purification system characterized in that it has, all samples with forward and reverse primer directly order deciding analysis against the NIAS ABI3730 XL genetic analysis machine have been performed. Direct order deciding of the code conditions based upon the total volume of 10 μl (sequencing buffer 5 μl, 1. 6 pM primer, 50 ng DNA of the genome, 0. 5 μl distilled water above and termination big die) 94 °C/10 seconds on and 60 °C/4 minutes as cycle, total 35 cycles have been performed. The reference the resultant product for 45 minutes 2,800 rpm into ethanol 70% 78% purification procedure to repel the cyclone comprises isocyanate. 2 hours at room temperature then drying the sample, are removed a formamide (10 μl) was added. The alignment of each sequence all DNAstar version 6. 0 of SeqMan program has executed, the genotype of a reflection sequences have been determined according to the apex. Dielectric inserted transformed for analysis, 1. 5% of agar gel electrophoretic system of copper executed, genotype have rapid mobility (276 bp, defective) with slow mobility (323bp, insertion) for DNA based on band pattern have been determined (also 1). Embodiments 5: statistical analysis Genotype general linear model (GLM) study influence conductor trait hosts analysis has been performed using statistical analysis system (SAS, 1990) procedure. Least squares mean is 0. 05 has been the comparison using a comparison of Fischer minimum consideration official approval error rate. Gene between two homozygous genetic type solution for home effect can be assessed by the other point, heterozygous genetic type solution otherwise dominant variation between two homozygous genetic type solution it has been determined that by means. Minimum mean square error of the markers for each standard has been measured by use of the following linear model: Y=μ + G + e, traits for observing value and Y is, μ is the total average and each trait, genotype G is fixed effect and, residual error e is are disclosed. Allele frequencies of Hardy - the [wey[wey] is equilibrium is bug Arlenquin version 3. 0 it has been determined that using. Haploview for analysis using software of an associated unbalanced were measured. Experiment result SUMMARY technical transformation to table 1 where exhibits. Amplification is a fragment of the genome of 267 and 323 bp has been generated. The analysis of the present invention decided according to genotype mobility of DNA band pattern. I.e., rapid DNA band (267 bp) on slow DNA band (323 bp) I (insertion) has been assigned to each D alleles (defective). In addition, sequencing is trapped in nucleotide position 81966364 81966429 56bp between insertion of his basis according to the sequence. PCR analysis of the present invention having two or more 2 insertion of any animal also found over yet, 195 (about 10%) have been cattle embryo the insertion of fragments. On the determination of the frequency alleles for each D I 0. 844 and 0. 156 (also 1) has been determined. In the alignment of the nucleotide sequence in the promoter regions of 5 81966235 81966377 (C>T) and (T>C) demonstrated that substitutions. G. 81966235C>T SNP is in C (0. 828) and T (0. 172) allele frequency race data, i.e., CC is 68. 5%, the TC 28. 5%, 3% TC is occupy other. CC, CT and TT genotype frequency for each 47. 1%, 43. 1% and 9. 8% of the first, g. 81966377T>C G for (0. 686) and A (0. 314) allele frequency's desire. G. 81966364D>I in SNP is D (0. 844) and I (0. 156) allele frequency show, DD is 71. 2%, the DI 26. 5%, the II 2. 4% of the first substrate. The significant deviation from Hardy - the [wey[wey] is SNP for retrieving air pressure of from measurement of the insert not found yet. Haplotype analysis insertion (g. 81966364D>I) and g. 81966377T>C significant between associated unbalanced has been confirmed. SNP ADIPOQ gene association rules between the result of the analysis to table 2 in the field of view of vehicle from the outside. G. 81966337T>C dielectric effect on the MAR LEA (P<0. 05) and on CAW BFT (P<0. 01) significance phenotypic difference are described. CC genotype (6. 054) and TC (5. 944) animals having TT genotype (5. 130) than with animal has significantly higher MAR too soon, of the genes set forth phase effect significant purges. While, opposite patterns are observed but, having animals alleles C T alleles having higher than animals BFT, CAW LEA and value of purges. Analysis is also significant recognition while LEA only home-effect, the basis for his home-and dominant effect and CAW BFT. G. 8196235C>T and g. 81966364D>I main alleles (major allele) genotype (alleles C and D (defective)) is lower than the value MAR ratings show different alleles, the analysis can be dominant (g. 8196235C>T) and move (g. 81966364D>I) effect was the basis. In addition to other markers for MAR significant association identifies a g. 8196235C>T and g. 81966364D>I (table 2) not found in his. Content of at least a portion shaped conductor that are directionally specific techniques, having conventional knowledge in the party industry to, such procedure only preferred embodiments only and in particular, the scope of this invention are inhibited will apparently not point. If substantial range defined by appended claim and their equivalent that will. The present invention relates to a marker for predicting a marbling score in hanwoo (Korean cattle) and a method for diagnosing the same. More particularly, the present invention relates to a marker for predicting a marbling score in hanwoo capable of detecting insertion or deletion in the base at position 258 of a polynucleotide derived from ADIPOQ gene expressed by SEQ ID NO: 1, or a marker for predicting a marbling score in hanwoo capable of detecting single base polymorphism in which the base at position 73 of a polynucleotide derived from ADIPOQ gene expressed by SEQ ID NO: 1 is C or T, a composition, a kit, and a method for predicting a marbling score in hanwoo by using the gene marker for predicting a marbling score in hanwoo. By applying the gene marker according to the present invention, a marbling score in hanwoo can be predicted in an effective and practical method. Also, based on the present invention, it is possible to select a seed bull which produces high quality of hanwoo meat. The sequence numbers 1 gene can be used for immobilizing polynucleotide can be inserted or in detecting defects in ADIPOQ derived from cattle e [pul[pul] ring 258 microarray for predicting polynucleotide. The cattle e [pul[pul] ring comprising a polynucleotide according to Claim 1 for predicting composition. According to Claim 2, 2 and 3 of the polynucleotide sequence number primer set compositions. According to Claim 3 composition kit for predicting e [pul[pul] ring 2 first anti or cattle. (A) extracting DNA genome of cattle; (b) the step (a) extracted from the genome DNA as templates, the sequence numbers 2 and 3 an amplifying genomic DNA for primer set mold step; and (c) amplifying the consequences to, sequence number 1 258 of second base sequence represented by sequence number 4 is not performed relative to plate is inserted polynucleotides occurs rating determining method for predicting good cattle e [pul[pul] ringe [pul[pul] ring cattle. The sequence numbers 1 gene can be used for immobilizing polynucleotide derived from a second base is T or C ADIPOQ 73 can be detecting a single nucleotide polymorphism in cattle for predicting e [pul[pul] ring polynucleotide. The cattle e [pul[pul] ring comprising a polynucleotide according to Claim 6 for predicting composition. According to Claim 7, 2 and 3 of the polynucleotide sequence number primer set compositions. 7 according to Claim 8 composition kit for predicting e [pul[pul] ring first anti or cattle. (A) extracting DNA genome of cattle; (b) the step (a) extracted from the genome DNA as templates, the sequence numbers 2 and 3 an amplifying genomic DNA for primer set mold step; and (c) amplifying the consequences to, sequence number 1 73 of second nucleotide sequence allele type is more excellent than when CT or TT CC e [pul[pul] ring grade cattle when determining method for predicting e [pul[pul] ring cattle. (A) extracting DNA genome of cattle; (b) the step (a) extracted from the genome DNA as templates, the sequence numbers 2 and 3 an amplifying genomic DNA for primer set mold step; and (c) amplifying the consequences to, sequence number 1 258 of second base sequence represented by sequence number 4 has taken place is inserted polynucleotides, sequence number of CT-allele 1 73 when a second nucleotide sequence, the sequence numbers 1 258 of second base sequence represented by polynucleotides is inserted sequence number 4 does not occur and, second nucleotide sequence allele type 73 when determining further excellent rating e [pul[pul] ring CC or TT cattle than cattle e [pul[pul] ring prediction method.
