KR101823376B1 - SNP marker regulating stearic acid level in the pork and uses thereof - Google Patents

Abstract

The present invention relates to a composition for determining a stearic acid content of pork containing a substance for detecting or amplifying SNP associated with a stearic acid content of pork, a kit for determining a stearic acid content of pork including the composition, a method of determining a stearic acid content of pork, a method of manufacturing pork with an increased stearic acid content, a method of increasing the stearic acid content of pork, and a marker composition and microarray for determining the stearic acid content of pork. Since an SNP marker for determining a stearic acid content of pork is used to objectively evaluate the stearic acid content of pork, which is not able to be distinguished visually, the present invention is able to be widely used for determining the quality of pork.

Description

SNP markers for regulating the content of stearic acid in pork and uses thereof Field of the Invention < RTI ID = 0.0 >

The present invention relates to a SNP marker for controlling the content of stearic acid in pork and a use thereof. More particularly, the present invention relates to a method for detecting a content of stearic acid in a pork containing an agent capable of detecting or amplifying a SNP associated with the content of stearic acid in pork A method for determining the content of stearic acid in pork, a method for producing a porcine with increased content of stearic acid in pork, a method for increasing the content of stearic acid in pork, a method for determining the content of stearic acid in pork, A marker composition for determining the content of stearic acid in pork, and a microarray.

Since breeding in the eastern part of India about 9,000 years ago, pigs have been raised as the most basic animal for the worldwide consumption of protein that people need, according to their age, situation and people's preferences. European varieties and Asian varieties are derived from Susscrofa on each continent. There are currently 200 varieties of varieties presently available. According to a recent FAO (Finance and Accounts Office) report, Asian cultivars account for 30% , And European varieties were reported to be around 33%. Differences in phenotype among these varieties are sought, size, and body shape.

Recently, in order to improve the quality of the pork meat, the pork is raised in a certain standard and is scientifically managed, and the pork meat obtained from the pork meat is branded, and various brands of pork meat are already commercially sold. However, since the branded pork meat and unbranded pork meat are difficult to identify with the naked eye, they can be used to sell the unbranded pork meat into branded pork meat, Disturbing events are occurring frequently. Indeed, since it is very difficult to distinguish between branded pigs and unbranded pork meat at the level of experts, studies are actively being conducted to establish objective criteria to judge the quality of genuine pork meat.

As a part of this research, a method for judging pig meat of a genuine brand was developed by analyzing the gene of pork meat. Randomly amplified polymorphic DNA (RAPD), single strand conformation polymorphisms, and other DNA analysis techniques to develop and develop a method for discriminating pork meat varieties. For example, Japanese Patent Application Laid-Open No. 2004-0039059 discloses a gene assay method capable of selecting pigs having excellent traits of pigs by using specific DNA markers related to the daily gain of body weight, backfat thickness, and meat quality traits of pigs. Patent Publication No. 2007-0113336 discloses a DNA marker for confirming the increase of porcine myocyte count using a mutation (SNP) caused by a single base sequence difference in the promoter region of the 5 'end of Myogenin gene known to be involved in the myocyte differentiation of pigs Japanese Patent Application Laid-Open No. 2011-0011443 discloses a technique for detecting an accurate Korean rice variety and other breeder pigs by detecting a Korean native pig-specific DNA marker for the KIT gene, -0050261 discloses a method of discriminating the varieties of black-bred pigs using haplotypes estimated from SNPs in the KIT gene region of black-bred pigs Patent Publication No. 2011-0139011 discloses a method for evaluating meat quality using a single-trait polymorphic biomarker for diagnosing fat content in pigs. In Patent Publication No. 2012-0046968, And a method of screening pigs of a trait of good quality using a gene of a pig is disclosed in Patent Publication No. 2012-0049624, (SNP) site of the PPARGC1A gene, which is involved in the meat quality of the pig, has been disclosed. In the patent publication No. 2012-0072871, there is disclosed a single gene for confirming the content of unsaturated fatty acids in pigs A method for identifying high quality pork meat using a base polymorphism (SNP) marker is disclosed.

On the other hand, stearic acid is a saturated fatty acid exhibiting various functionalities and is known to affect the meat quality of meat. The content of stearic acid in pork can be used as a standard for judging the quality of pigs, A method for evaluating the content of stearic acid has not yet been developed.

Under these circumstances, the present inventors have made extensive efforts to develop a method for evaluating the content of stearic acid in pork using a gene. As a result, it has been confirmed that the content of stearic acid in pork can be evaluated by using SNPs present on chromosome 14 And completed the present invention.

One object of the present invention is to provide a composition for determining the content of stearic acid in pork comprising a preparation capable of detecting or amplifying the SNP associated with the content of stearic acid in pork.

Another object of the present invention is to provide a kit for determining the content of stearic acid in pork comprising the above composition.

Another object of the present invention is to provide a method for determining the content of stearic acid in pork using the above composition.

Yet another object of the present invention is to provide a method for producing pigs with increased content of stearic acid in pork.

It is another object of the present invention to provide a method for increasing the content of stearic acid in pork.

Yet another object of the present invention is to provide a marker composition for determining the content of stearic acid in pork.

Yet another object of the present invention is to provide a microarray for determining the content of stearic acid in pork containing the marker.

The present inventors searched quantitative trait loci (QTL) related to saturated and unsaturated fatty acid contents of pigs in order to develop a method for reducing the content of saturated fatty acids in pigs. As a result, SNPs located on chromosome 14 H3GA0042070) is associated with the content of stearic acid in pork, and it can be used as a genetic marker which can determine the content of stearic acid in pork using the SNP. The technique of using SNP (H3GA0042070) located on chromosome 14 to determine the content of stearic acid in pork has not been known until now and was developed by the present inventor for the first time.

In one embodiment, the present invention provides a polynucleotide comprising the nucleotide sequence of SEQ ID NO: 1, wherein the base 501 is T or C, and the single nucleotide polymorphism (SNP) located at the base 501 is included A composition capable of detecting or amplifying a polynucleotide composed of 5 to 1000 consecutive bases or a complementary polynucleotide thereof, which is capable of detecting or amplifying a stearic acid content in pork.

The term "polynucleotide consisting of the nucleotide sequence of SEQ ID NO: 1" of the present invention is a polymorphic sequence comprising a polymorphic site of a gene involved in the content of stearic acid in pork, wherein the polymorphic sequence is a polynucleotide sequence Quot; means a sequence comprising a polymorphic site comprising a SNP. The polynucleotide sequence may be DNA or RNA.

The term "polymorphism " of the present invention means a case where two or more alleles exist in one locus, and among polymorphic sites, only one single base is different from the single It is called single nucleotide polymorphism (SNP). Specific polymorphic markers have two or more alleles that exhibit an incidence of 1% or more, more preferably 5% or 10% or more in the selected population.

The term "allele " of the present invention refers to various types of a gene existing on the same locus of a homologous chromosome. Alleles are also used to represent polymorphisms, for example, SNPs have two kinds of bialles.

Terms of this invention "stearic acid (stearic acid) (C18: 0 )" is, is represented by the chemical formula of C ₁₈ H ₃₆ O ₂ with 18 carbon atoms, the melting point and any point of about 69.4 ℃ of about 71 ℃ ≪ / RTI >

In the present invention, the stearic acid is used as an object capable of predicting the content of pork using the genotype of the SNP (H3GA0042070) located on the chromosome 14. The pork containing the stearic acid is not particularly limited, Can be pork derived from native pigs, land races or hybrids of these pigs.

The term "agent capable of detecting or amplifying a polynucleotide" of the present invention refers to an agent capable of specifically recognizing binding to a polymorphic site containing a SNP in a polynucleotide and amplifying the polymorphic site As a preparation, specifically, a probe capable of specifically binding to a polymorphic site containing a SNP, a polynucleotide comprising the SNP marker, or a primer capable of specifically amplifying a complementary polynucleotide of the polynucleotide may be used.

In the present invention, a probe used to recognize and bind to a SNP marker includes a sequence complementary to a polynucleotide sequence including a SNP, and may be a DNA, RNA, or DNA-RNA hybrid form . Further, fluorescent markers, radiation markers, and the like can be additionally attached to the 5 'or 3' ends of the probe so as to be visually recognizable.

The term "primer " of the present invention refers to a nucleotide sequence having a short free 3 'hydroxyl group, capable of forming a base pair with a complementary template, Means a short sequence functioning as a starting point. The primers used in the present invention for the amplification of SNP markers can be amplified by PCR using appropriate conditions in suitable buffers (for example, 4 different nucleoside triphosphates and polymerase such as DNA, RNA polymerase or reverse transcriptase) Stranded oligonucleotide that can serve as a starting point for the directed DNA synthesis. The appropriate length of the primer may vary depending on the purpose of use, but it may be generally used in a size of 15 to 30 nucleotides. The primer sequence is not necessarily completely complementary to the polynucleotide comprising the SNP marker or its complementary polynucleotide, and can be used if it is sufficiently complementary to hybridize.

The primers can also be modified, for example, by methylation, capping, substitution of nucleotides or modifications between nucleotides such as uncharged linkers (e.g., methylphosphonate, phosphotriester, phosphoramidate, Carbamates, etc.) or charged linkages (e.g., phosphorothioates, phosphorodithioates, etc.). As an example, the primer may be a primer base pair consisting of SEQ ID NOS: 2 and 3 capable of amplifying a polynucleotide consisting of the nucleotide sequence of SEQ ID NO: 1, but is not limited thereto.

In the present invention, the content of stearic acid in the pork is the polynucleotide comprising the nucleotide sequence of SEQ ID NO: 1, wherein the alleles of base No. 501 are all C, Can be judged to contain stearic acid in a higher content than the pork derived from pork.

According to one embodiment of the present invention, in order to investigate the fatty acid content, the fattening of the fattened shrimp was sampled for the offspring of the Korean traditional pig and the land race reference group F2, The content of stearic acid was found to be the third highest in the total fatty acids, and the content thereof was 13.65% (FIG. 1). In addition, analysis of the loci of pigs related to stearic acid in pork confirmed that the SNP (H3GA0042070) located on chromosome 14 was associated with the content of stearic acid in pork (Fig. 2 and Table 1) Analysis of the content of stearic acid in pork showed that the content of stearic acid was the highest when the genotype of the SNP was CC and the content of stearic acid was relatively high when the genotype was TT And the content of stearic acid was a median value when the genotype was CT (Table 2).

Thus, it can be seen that the SNP of SEQ ID NO: 1 can be used as a gene marker for determining the content of stearic acid in pork.

As another embodiment for achieving the above object, the present invention provides a kit for determining the content of stearic acid in pork comprising the composition. Specifically, the kit may be a PCR (Polymerase Chain Reaction) kit or a DNA analysis kit (for example, a DNA chip).

The kit of the present invention can determine the content of stearic acid in pork by confirming the genotype of the SNP marker provided by the present invention using the above composition by amplification or by checking the expression level of mRNA. The kit provided in the present invention may be a kit containing essential elements necessary for performing RT-PCR.

For example, in addition to the respective primer pairs specific for the SNP for determining the content of stearic acid in the pork, the RT-PCR kit can also be used in test tubes or other appropriate containers, reaction buffers (pH and magnesium concentrations vary ), Deoxynucleotides (dNTPs), enzymes such as Taq polymerase and reverse transcriptase, DNase, RNAse inhibitors, DEPC-water, sterile water, and the like. It may also contain a primer pair specific for the gene used as a quantitative control.

As another example, the kit of the present invention may be a DNA chip kit for determining the content of stearic acid in pork containing elements necessary for carrying out a DNA chip.

The term "DNA chip" of the present invention means one of DNA microarrays capable of confirming each base of hundreds of thousands of DNAs at a time.

The DNA chip kit is formed by attaching nucleic acid species to a glass surface, which is generally not larger than a flat solid support plate, typically a slide for a microscope, in a gridded array. The nucleic acid is uniformly arranged on the chip surface, It is a tool that enables multiple parallel hybridization reactions between the nucleic acid on the chip and the complementary nucleic acid contained in the treated solution on the chip surface.

(A) a polynucleotide comprising the nucleotide sequence of SEQ ID NO: 1 or a complementary nucleotide thereof, from the DNA of the sample separated from the individual, Amplifying; And (b) determining the base of the amplified polymorphic site. At this time, DNA of the separated sample can be obtained from a sample isolated from an individual.

The term "individual" in the present invention means a pig to which the content of stearic acid in pork is to be confirmed. By analyzing the genotype of the polymorphic site of the SNP using the sample obtained from the pig, The content of stearic acid can be judged. Examples of the specimen include, but are not limited to, hair, urine, blood, various body fluids, separated tissues, separated cells or saliva, and the like.

The step of amplifying the polymorphic site of the SNP contained in the marker from the DNA of step (a) can be used in any method known to a person skilled in the art. For example, the target nucleic acid can be obtained by PCR amplification and purification thereof. Other ligase chain reaction (LCR) (Wu and Wallace, Genomics 4, 560 (1989), Landegren et al., Science 241, 1077 (1988)), transcription amplification (Kwoh et al., Proc. Natl. Acad. Sequence amplification based on nucleic acids (NASBA) can be used as well as self-sustaining sequence replication (Guatelli et al., Proc. Natl. Acad. Sci. USA 87, 1874 (1990)).

Determination of the base of the amplified polymorphic site in step (b) of the above method can be carried out by sequencing, hybridization by microarray, allele specific PCR, dynamic allelespecifichybridization, DASH), PCR extension analysis, PCR-SSCP, PCR-RFLP analysis or TaqMan technique, SNPlex platform (Applied Biosystems), mass spectrometry (e.g. MassenRAY system of Sequenom), mini- (E.g., BioRad), CEQ and SNPstream system (Beckman), Molecular Inversion Probe array technology (e.g. Affymetrix GeneChip), and BeadArray Technologies (e.g. Illumina GoldenGate and Infinium assay) But is not particularly limited thereto. One or more alleles in a SNP marker comprising the mutation site can be identified by the methods described above or other methods available to those skilled in the art to which the invention pertains. Determination of the base at such a mutation site can be carried out specifically through a DNA chip.

The TaqMan method comprises the steps of (1) designing and preparing a primer and a TaqMan probe to amplify a desired DNA fragment; (2) labeling probes of different alleles with FAM dyes and VIC dyes (Applied Biosystems); (3) performing PCR using the DNA as a template and using the primer and the probe; (4) after completion of the PCR reaction, analyzing and confirming the TaqMan assay plate with a nucleic acid analyzer; And (5) determining the genotype of the polynucleotides of step (1) from the analysis results.

The sequencing analysis can be performed using a conventional method for nucleotide sequencing, and can be performed using an automated gene analyzer. In addition, allele-specific PCR means a PCR method in which a DNA fragment in which the mutation is located is amplified with a primer set including a primer designed with the base at the 3 'end at which the mutation site is located. The principle of the above method is that, for example, when a specific base is substituted with C in T, an opposite primer capable of amplifying a primer containing the T as a 3 'terminal base and a DNA fragment of an appropriate size is designed and PCR In the case of performing the reaction, when the base at the mutation position is T, the amplification reaction is normally performed and a band at a desired position is observed. When the base is substituted with C, the primer can be complementarily bound to the template DNA, And the amplification reaction is not performed properly due to the inability of complementary binding at the terminal. DASH can be carried out by a conventional method, specifically, by the method by Prince et al.

On the other hand, in the PCR extension analysis, first, a DNA fragment containing a base in which a single nucleotide polymorphism is located is amplified with a pair of primers, and all the nucleotides added to the reaction are deactivated by dephosphorylation, and a specific extension primer, dNTP And then performing a primer extension reaction by adding a mixture, a digoxinucleotide, a reaction buffer, and a DNA polymerase. At this time, the extension primer has the 3 'end immediately adjacent to the 5' direction of the base where the mutation site is located, and the nucleic acid having the same base as the didyoxynucleotide is excluded in the dNTP mixture, and the didyoxynucleotide has a mutation ≪ / RTI > For example, when dTTP, dCTP, and TTP mixture and ddATP are added to the reaction in the presence of a substitution from T to C, the primer in the substituted base is extended by DNA polymerase, and after a few bases, T The primer extension reaction is terminated by ddATP at the position where the base first appears. If the substitution has not occurred, the extension reaction is terminated at the position, so that it is possible to discriminate the kind of base showing the mutation by comparing the length of the extended primer.

At this time, as a detection method, when the extension primer or the didyxin nucleotide is fluorescently labeled, the mutation is detected by detecting fluorescence using a gene analyzer (for example, Model 3700 manufactured by ABI Co., Ltd.) used for general nucleotide sequence determination And when the unlabeled extension primer and the didyxin nucleotide are used, the genetic variation of the SNP can be detected by measuring the molecular weight using MALDITOF (matrix assisted laser desorption ionization-time of flight) technique.

Specifically, the pork pork in which the allele of the polynucleotide consisting of the nucleotide sequence of SEQ ID NO: 1 in the base sequence determined in the step (b) is all the C-501 allele is the pork pork of all the T alleles It can be judged that the content of stearic acid is increased.

According to another aspect of the present invention, there is provided a method of isolating a SNP trait that is involved in the content of stearic acid in pork, comprising the step of contacting a polynucleotide consisting of the nucleotide sequence of SEQ ID NO: And a step of immobilizing all of the genes to C, wherein the content of stearic acid in the pork is increased.

According to another aspect of the present invention, there is provided a method of isolating SNPs of genes involved in the content of stearic acid in pork, comprising the steps of: isolating a polynucleotide consisting of the nucleotide sequence of SEQ ID NO: 1 Wherein all the alleles of the pancreas are immobilized in C. The present invention also provides a method for increasing the content of stearic acid in pork.

Specifically, the step of immobilizing the SNP trait may be carried out by crossing an individual having the SNP trait and selecting an individual having the desired trait. More specifically, a polynucleotide consisting of the nucleotide sequence of SEQ ID NO: 1, which is involved in the content of stearic acid in pork, is crossed with an individual having C / C at the 501st base, and the polynucleotide of SEQ ID NO: 1 By selecting pigs fixed with C / C at base number 501, pigs with increased content of stearic acid in pork can be obtained. The pig may be a hybrid of Jeju native pig, land lace or Jeju native pig and land lace.

In another aspect of the present invention, there is provided a marker composition for determining the content of stearic acid in pork, which comprises a polynucleotide comprising the nucleotide sequence of SEQ ID NO: 1.

In the present invention, using the marker, the pork pork in which the allele of the polynucleotide consisting of the nucleotide sequence of SEQ ID NO: 1 is all of base number 501 is selected from the group consisting of stearic acid Of the total amount of water.

In another aspect of the present invention, there is provided a microarray for determining the content of stearic acid in pork comprising the marker composition for determining the content of stearic acid in the pork.

The microarray may comprise DNA or RNA polynucleotides. The microarray comprises a conventional microarray except that the polynucleotide of the present invention is contained in the probe polynucleotide.

Methods for producing microarrays by immobilizing probe polynucleotides on a substrate are well known in the art. The probe polynucleotide means a polynucleotide capable of hybridizing, and means an oligonucleotide capable of binding to the complementary strand of the nucleic acid in a sequence-specific manner. The probe of the present invention is an allele-specific probe in which a polymorphic site exists in a nucleic acid fragment derived from two members of the same species and hybridizes to a DNA fragment derived from one member but does not hybridize to a fragment derived from another member . In this case, the hybridization conditions show a significant difference in the intensity of hybridization between alleles, and should be sufficiently stringent to hybridize to only one of the alleles. This can lead to good hybridization differences between different allelic forms. The probe of the present invention can be used for determining the content of stearic acid in pork by detecting an allele. The determination method includes detection methods based on hybridization of nucleic acids such as Southern blot, and may be provided in a form pre-bonded to a substrate of a DNA chip in a method using a DNA chip. The hybridization can usually be performed under stringent conditions, for example, a salt concentration of 1 M or less and a temperature of 25 ° C or higher. For example, conditions of 5x SSPE (750 mM NaCl, 50 mM Na Phosphate, 5 mM EDTA, pH 7.4) and 25-30 < 0 > C may be suitable for allele-specific probe hybridization.

The process of immobilizing the probe polynucleotide on the substrate in connection with the determination of the content of stearic acid in the pork of the present invention can also be easily carried out using this conventional technique. In addition, hybridization of nucleic acids on a microarray and detection of hybridization results are well known in the art. The detection can be accomplished, for example, by labeling the nucleic acid sample with a labeling substance capable of generating a detectable signal comprising a fluorescent material, such as Cy3 and Cy5, and then hybridizing on the microarray and generating The hybridization result can be detected.

Since the SNP marker capable of determining the content of stearic acid in the pork of the present invention is used as a means for objectively evaluating the content of stearic acid in the pork which is not visually distinguishable from the naked eye, will be.

1 is a schematic view showing the composition of fatty acids in pork;
FIG. 2 is a graph showing the result of analyzing a genome wide association study (GWAS) using a DNA chip as a chromosome containing a gene related to the content of stearic acid in pork.
Fig. 3 is a schematic diagram showing various gene groups in the locus of the content of stearic acid in the pork of chromosome 14.
FIG. 4 is a graph showing the result of identifying the genotype of the 501 (C / T) SNP contained in the polynucleotide of SEQ ID NO: 1 of chromosome 14 using pyrosequencing.

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  One: In pork  Fatty acid composition analysis

In order to analyze the composition of saturated and unsaturated fatty acids in pigs, the content of fatty acid composition was investigated in about 1,000 dogs in the second generation offspring of Jeju traditional pig and landrace mating reference group.

1 is a schematic view showing the composition of fatty acids in pork; As shown in FIG. 1, the content of stearic acid was 13.65%, which is the third highest content of total fatty acids.

Example  2: In pork Stearic acid  Content association Gene locus  Search

In order to analyze the gene region regulating the content of stearic acid in pork, GWAS was analyzed with a large DNA chip (illumina 60k SNPchip) to search for the locus of saturated fatty acid content-related gene (Fig. 2).

FIG. 2 is a graph showing the result of analyzing a genome wide association study (GWAS) using a DNA chip as a chromosome containing a gene related to the content of stearic acid in pork. As shown in FIG. 2, it was confirmed that the locus associated with the content of stearic acid in the pork had the highest significance on the chromosome 14 of the pig.

Example  3: In pork Stearic acid  Securing content-related SNP

As a result of confirming that the content of stearic acid in the pork was determined by the locus corresponding to the chromosome 14 through the above Example 2, SNPs highly correlated with the content of stearic acid in the pork were selected (Table 1).

SNPs that are highly related to stearic acid content in pork chromosome SNP BP LOG 14
14
14
14
14
14 H3GA0042070
ALGA0081025
SCD_1-2
ALGA0081396
ALGA0081481
ALGA0081112 121721589
119955763
120964624
129795414
131807843
121527590 24.01390037
23.46864884
23.08465294
18.74763249
16.88239731
16.06429125

As shown in Table 1, the highest SNP associated with the content of stearic acid in the pork of chromosome 14 was H3GA0042070.

Example  4: In pork Stearic acid  Content Chromosome 14 Gene locus  Identification of Genes in the Region

Based on the SNP list having high content of stearic acid content in the pork of Example 3, various gene groups in the gene locus region of chromosome 14 associated with the content of stearic acid in pork were confirmed (Fig. 3).

Fig. 3 is a schematic diagram showing various gene groups in the locus of the content of stearic acid in the pork of chromosome 14. As shown in FIG. 3, the genes related to the content of stearic acid in the pork chromosome 14 contained CNNM1, NKX2-3, COX15, CPN1, SNORA12, HIF1AN, PAX2, SEMA4G and MRPL43.

Example  5: Pyro-sequencing ( Phyro -sequencing In pork Stearic acid  Extraction and analysis of nucleotide sequences containing content-related SNPs

Through the above Example 3, it was confirmed that H3GA0042070 of chromosome 14 had the highest significance with respect to the content of stearic acid in pork, and 1001 nucleotides (SEQ ID NO: 1) were respectively ensured around each SNP present on chromosome 14 And their nucleotide sequences were analyzed.

In order to amplify the polynucleotide of SEQ ID NO: 1 of chromosome 14, PCR was carried out using the DNA obtained from a number of pigs as a template and the following primers to obtain a polynucleotide fragment of SEQ ID NO: 1 of chromosome 14.

Forward: 5'-AGG-AGT-TTG-GGG-AGG-CTT-T-3 '(SEQ ID NO: 2)

Reverse: 5'-biotin-GCC-CAT-TAA-GGC-AAT-TGA-GA-3 '(SEQ ID NO: 3)

The nucleotide sequence of the polynucleotide fragment of SEQ ID NO: 1 obtained using the above primer was determined and the genotype of the SNP contained therein was confirmed.

In order to confirm the 501 (C / T) SNP, the nucleotide sequence of the polynucleotide fragment was determined using the following primer (Mini-seq), and the primer of the input was used. / T) SNP genotype (Fig. 6).

GGG-TGA-CCC-TGG-GA-3 '(SEQ ID NO: 4)

Input: 5 '- [C / T] TTGGGGTTGCT-3' (SEQ ID NO: 5)

FIG. 4 is a graph showing the result of identifying the genotype of the 501 (C / T) SNP contained in the polynucleotide of SEQ ID NO: 1 of chromosome 14 using pyrosequencing. As shown in FIG. 4, it was confirmed that polynucleotides of amplified SEQ ID NO: 1 contained TT, TC or CC genotypes, respectively.

Example 6: 14  Genotype of SNP of chromosome In pork Stearic acid  Analysis of association with content

The changes in the content of stearic acid in the pork according to the genotype of chromosome 14 SNP identified in Example 5 were compared (Tables 2 and 3).

Analysis of the relationship between the genotype of the chromosome 14 SNP in pig and the content of stearic acid in pork genotype Analysis Stearic acid content TT
CT
CC 551
378
23 13.6688 ± 1.06
14.0705 + - 1.12
14.3748 ± 1.12

As shown in Table 2, when the genotype of chromosome 14 SNP was CC, the content of stearic acid in the pork was the highest, and when the genotype was TT, the content of stearic acid in the pork was the lowest And the content of stearic acid in the pork was a medium level when the genotype was CT.

<110> REPUBLIC OF KOREA (MANAGEMENT: RURAL DEVELOPMENT ADMINISTRATION) <120> SNP marker regulating stearic acid levels in the pork and uses          the <130> KPA160577-KR <160> 5 <170> Kopatentin 2.0 <210> 1 <211> 1001 <212> DNA <213> Artificial Sequence <220> <223> SNP <400> 1 aaagtgagac tgatttaaaa gtgttcttat gaggattaat gaactgatgg gtataacaca 60 taaaagcttt aaagccccct acaaatcagg gtcgttataa ccacttgccc caccctcagt 120 gtgaaaccaa attgtacctt gaagctcaca cttcacacag catctggatc ctgccctgat 180 tcccatccca gcaagaatat cgttaaggac atagaaagct tgatgaagag ctgttaaatg 240 aaaaattgaa gggatggaac acagtctaat ctctggtctc tgtgaagagg ccatactggt 300 ggagggtttg ggggagcaac gtagggaaga gagaaaactg gagtcagaac actggccagg 360 agtttgggga ggctttgggg gagtgggaaa aagtgggttc agggcatctg ggggtctctg 420 aagcagagtt tgtggtgcac agagggaaag tttgaggttg ggtgctgagg caagatgtgg 480 ggctgggt ggcaggtgga agatgggggt ccggccaacc caagtctctc tctgtccggc aggaggggac 600 tctggtgcag ggcgaggagg gagattagga ggcgctctca attgccttaa tgggctctaa 660 gaggcgggat cctattacag ctcggggtgc cccggggtgc cttgctggga agacaagcta 720 atctgccatg caaattaaat cctgcctcaa gacgcaaggc ctgcttaatg tgatccatta 780 cacagtttac ttgtttatat ctaatattgg aacaggcagc ccgggcgggc ggcagcggca 840 ggcggagcgg cccaaaagca ggagagtaaa ttaccagcaa aatgagcttt tccatcctta 900 gccgccgcca cccgcgcctt ggtggctctc tgtgtcctgg gcgcccccgc ccgcccgccc 960 tcccgcagcg tcccctcgct ctctgggctg tcccgctctc t 1001 <210> 2 <211> 19 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 2 aggagtttgg ggaggcttt 19 <210> 3 <211> 20 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 3 gcccattaag gcaattgaga 20 <210> 4 <211> 20 <212> DNA <213> Artificial Sequence <220> <223> mini-seq <400> 4 aaaagggggt gaccctggga 20 <210> 5 <211> 12 <212> DNA <213> Artificial Sequence <220> <223> input seq <400> 5 yttggggttg ct 12

Claims (12)

A polynucleotide consisting of 5 to 1000 consecutive bases comprising a single nucleotide polymorphism (SNP) located at the 501 base, wherein the polynucleotide consisting of the nucleotide sequence of SEQ ID NO: 1 is T or C at position 501, A composition for judging the content of stearic acid in pork comprising an agent capable of detecting or amplifying a complementary polynucleotide,
It is judged that the content of stearic acid in the pork having all the alleles of base No. 501 of the polynucleotide having the nucleotide sequence of SEQ ID NO: 1 is higher than the content of stearic acid in the pork having the alleles of T A composition for judging the content of stearic acid in pork.
The method according to claim 1,
Wherein the preparation comprises a primer pair consisting of SEQ ID NOS: 2 and 3 capable of amplifying a polynucleotide comprising the nucleotide sequence of SEQ ID NO: 1.
The method according to claim 1,
Wherein the formulation is used to determine the content of stearic acid in pork obtained from Jeju native pigs, land lace or Jeju native pigs and landrace hybrids.
A kit for determining the content of stearic acid in pork comprising the composition of any one of claims 1 to 3.
(a) amplifying a polynucleotide consisting of the nucleotide sequence of SEQ ID NO: 1 or a polymorphic site of the complementary polynucleotide of the polynucleotide of SEQ ID NO: 1 from the DNA of the sample isolated from the individual; And
(b) determining a base of the amplified polymorphic site, comprising the steps of:
It is judged that the content of stearic acid in the pork having all the alleles of base No. 501 of the polynucleotide having the nucleotide sequence of SEQ ID NO: 1 is higher than the content of stearic acid in the pork having the alleles of T In method.
delete The step of fixing the SNP trait involved in the content of stearic acid in the pork comprises the step of fixing the allele of the polynucleotide consisting of the nucleotide sequence of SEQ ID NO: A method for producing an excellent pig having an increased content.
8. The method of claim 7,
The step of immobilizing the SNP trait is carried out by crossing an individual having alleles of base No. 501 of a polynucleotide consisting of the nucleotide sequence of SEQ ID NO: 1, by selecting an individual having a desired trait .
The step of fixing the SNP trait of the gene involved in the content of stearic acid in the pork comprises the step of fixing the allele of the polynucleotide consisting of the nucleotide sequence of SEQ ID NO: A method for increasing the content of ricin acid.
10. The method of claim 9,
The step of immobilizing the SNP trait is carried out by crossing an individual having alleles of base No. 501 of a polynucleotide consisting of the nucleotide sequence of SEQ ID NO: 1, by selecting an individual having a desired trait .
1. A marker composition for determining the content of stearic acid in pork, wherein the polynucleotide consisting of the nucleotide sequence of SEQ ID NO: 1 is T or C at position 501,
The polynucleotide consisting of the nucleotide sequence of SEQ ID NO: 1 indicates that the content of stearic acid in the pork in which each allele of the 501 base is C is higher than the content of stearic acid in the pork in which each of the alleles is T A marker composition for determining the content of stearic acid in pork.
A microarray for determining the content of stearic acid in pork comprising the marker composition of claim 11.

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