KR20110067941A - Dna markers for detecting increase of muscle fiber type i within porcine muscle - Google Patents

Abstract

PURPOSE: A DNA marker for detecting increase of muscle fiber type I is provided to select an individual with improved meat quality. CONSTITUTION: A DNA marker for detecting increase of muscle fiber type I in the porcine muscle comprise 15-500 serial sequence containing 678th T base in a sequence of sequence number 1. A method for confirming the increase of muscle fiber type I comprises: a step of isolating genomic DNA from a pig; a step of amplifying a specific site containing 678th base of PPARGC1A gene exon 8; a step of treating the amplified DNA with AluI; and a step of comparing gene fragments with restriction fragment length polymorphism(RFLP).

Description

DNA markers for detecting increase of muscle fiber type I within porcine muscle}

The present invention relates to a DNA marker for identifying an increase in muscle fiber type I in pigs, more specifically, consisting of 15-500 consecutive sequences including the 678th T base in SEQ ID NO: 1. The present invention relates to a DNA marker for checking the increase in myofibrillar type I.

For a long time, pig breeding has been aimed at increasing feed efficiency and growth rate, and has been progressing towards lean pork production, which has reduced fat and other fats and increased red meat ratio (Brocks et al., 2000). At present, there have been significant developments in red meat production efficiency, but this is mainly interpreted as a result of the growth of muscle cells (Rehfeldt et al., 2000). It caused degradation (Cameron, 1990). Therefore, it is urgent to prepare an improvement system for improving the meat quality together with the existing red meat production capacity.

Understanding the growth and development of skeletal muscle is of great importance in terms of meat production and meat quality of livestock livestock (Karlsson et al., 1993). Muscles are composed of muscle fibers, and muscle production is determined by the number and size of muscle fibers. Red meat production increases through the number of muscle fibers determined before birth and by the growth of muscle fibers after birth (Rehfeldt and Fiedler, 1984). However, in the case of meat quality, it is influenced by the composition of muscle fibers rather than the number and size of muscle fibers (Ryu and Kim, 2005).

Muscle muscle fibers in pigs can be classified into three types according to the type of myosin-heavy chain responsible for ATPase activity (Brooke and Kaiser, 1970). Myofibrillar type I (slow-oxidative) slows down the metabolic rate and has a positive effect on meat quality at high content, on the contrary, type IIb (type IIb) In the case of fast-glycolytic (fast-glycolytic) properties due to the rapid post-metabolism rate is high because the meat content is pale flesh, pale tissue tissue sulky and juicy (pale, soft, exudative; PSE) increases the likelihood of being identified as abnormal (Klont et al., 1998). The other, type IIa, has a slow-glycolytic property that is intermediate to both. Myofibrillar muscle composition is determined according to these three types of myofibrils. Porcine longissimus dorsi is mostly white muscle type, and the proportion of type IIb accounts for about 80%. Research to increase the ratio is needed.

Muscle fiber properties, including muscle fiber composition, are strongly influenced by genetic factors. In particular, pigs have a high heritability of 37-58% for each myofibrillar type and a high genetic correlation with meat-related traits. Larzul et al., 1997). Marker Assisted Selection (MAS), a selection method using DNA markers, increases the accuracy of selection and increases the rate of improvement in livestock breeding and maximizes genetic improvement. have. Much of the variation in myofibril composition is caused by genetic factors, suggesting that MAS-based improvements can be very efficient.

However, myocyte characteristics are traits that can only be measured in the cartilage of the carcass, and cannot be measured in vivo. Therefore, for selection and improvement based on this, there is a need for a method capable of predicting the myocyte characteristics of the individual without slaughter. In this regard, the development of DNA markers is fundamentally required, and the development of candidate genes and the securing of genetic resources through gene structure analysis must be essential.

MyofD formation in mammals is determined during embryonic development and the MyoD gene family has been proposed as a gene involved in this (te Pas et al., 2000). The biochemical mechanisms that control myofibrillar composition have not been elucidated so far, but have been reported to depend on energy absorption and metabolism (Berchtold et al., 2000). The Perogsome proliferator-activated receptor-gamma coactivator-1 (PPARGC1A) gene is a precursor of PGC1 and is deeply involved in animal metabolism by deeply involved in adaptive thermogenesis and adipocyte differentiation (Puigserver et al., 1998). Recently, it has been recently mentioned as a candidate for meat quality in livestock (Erkens et al., 2006). Moreover, since this gene has been reported to play a major role in inducing myofiber type I formation in mice (Lin et al., 2002), we have found that the PPARGC1A gene is involved in porcine myofiber, in particular type I formation and associated with meat quality improvement. Was selected as a candidate gene.

In the present invention, the actual muscle fiber and meat quality was measured in the Yorkshire varieties of pigs, and the correlation analysis was performed based on the genotyping results at the Cys430Ser polymorphic locus of the PPARGC1A gene. Through this, muscle fiber composition type I augmentation at the corresponding locus of the gene of the present invention and a function related to meat quality improvement were identified, and its efficacy as a related DNA marker was verified.

Therefore, the main object of the present invention is to increase the ratio of myofibrillar type I in the myofibrillar composition of the PPARGC1A gene of pigs by increasing the ratio of myofibrillar type I DNA markers related to pig myofiber and SNP analysis kit. To provide.

According to an aspect of the present invention, the present invention comprises a DNA marker for confirming the increase in muscle fiber type I in pigs, consisting of 15 to 500 consecutive sequences including the 678th T base in the nucleotide sequence of SEQ ID NO: 1 to provide. The inventors found a single nucleotide polymorphism (SNP) associated with intramuscular muscle fiber composition in pigs in exon 8 of the PPARGC1A gene represented by SEQ ID NO: 1, and the SNP was associated with intramuscular muscle fiber composition and genotypes from pig individuals. Association was analyzed. The DNA marker is a part of the sequence of the PPARGC1A gene, which has recently been studied as a meat-related candidate gene for livestock, and includes a single nucleotide polymorphism (SNP) site in which a base is substituted with A or T at a specific site.

The SNP causes a Cys430Ser mutation of exon 8 of the PPARGC1A gene. In brief, the SNP is present in exon 8 of the PPARGC1A gene and the genetic code for encoding the 430th amino acid of the entire amino acid sequence of the PPARGC1A gene is AGC or TGC. Induces a mutation (Cys430Ser) to coat different amino acids with Ser or Cys, respectively.

The present invention identifies a SNP (A → T substitution) site that causes an increase in myofibrillar type I in the entire sequence of the PPARGC1A gene, and is a novel DNA marker for confirming whether the myofibrillar myofibrillar type I increases in pigs. It is about one use.

The PPARGC1A gene is known to be involved in energy metabolism in mammals, and various studies have been conducted as a candidate for meat quality in livestock, and in particular, it is known to play a major role in inducing muscle fiber type I formation in mice. . However, there is no known method for selecting an individual having improved muscle fiber type I by using SNPs of a specific region in the pig PPARGC1A gene and improving meat quality, and the inventors first disclosed the SNP related to this muscle fiber type I increase.

In the present invention, the DNA fragment is a DNA marker comprising the 678th T base, which is a new genetic variation (SNP site) of the PPARGC1A gene exon 8 related to pig muscle fiber composition. This genetic mutation site is a substitution of a base from A to T, and the difference in gene expression, resulting in an increase in the proportion of type I in the composition of muscle fibers, which affects meat quality. The size of the DNA fragment may be any fragment size including the SNP site, unless it is the full length of the entire gene, but preferably 15 to several hundred bases, more preferably 15 to 500 bases. Especially 15 to 30 base can be used as a probe (probe) or a primer for detecting the SNP, in the case of a base having a size larger than that in the PCR-RFLP method, as in the embodiment of the present invention It can be used to For example, as shown in Figure 1, the DNA fragment of 200 bp size (SEQ ID NO: 4) including the SNP site was used as a DNA marker.

In addition, in the present invention, the improvement of meat quality is characterized by including an increase in pH and a decrease in meat color. As in Example 2 of the present invention, the analysis of the correlation analysis of genotypes (AA, AT, TT) and myofiber and meat quality according to SNP, AT, TT genotype with T allele has muscle fiber composition with increased muscle fiber type I This confirmed that the improved meat quality.

In the present invention, "increase in muscle fiber type I" refers to a relatively high proportion of muscle fiber type I compared to muscle fiber type IIa, IIb in the muscle fiber composition of pigs according to individual genotypes (AA, AT, TT) due to SNP Means that. That is, the composition ratio of muscle fiber type I is increased compared to AA genotype individuals.

The “PCR-RFLP” method used in the present invention analyzes restriction fragment length polymorphism of gene fragments using PCR (Polymerase Chain Reaction) technology among various DNA analysis techniques for improving genetic characteristics or ability of livestock. In this technique, when restriction enzyme recognition sites exist at specific point mutation sites, the polymorphisms of DNAs varying in the length of fragments generated by restriction enzymes according to the genotype difference of each individual are compared. It is a faster and more accurate method of determining the desired genotype.

In the present invention, the meat properties of pigs are classified into grades of meat quality according to pH, meat color, intramuscular fat degree, fat color, texture, maturity, etc. Among them, pH and meat color are particularly important criteria for indicating meat quality of pork. Becomes Since the price of pork is soaring recently, it is important to develop high quality meat with improved quality by selecting, breeding and producing pigs with excellent meat quality. Therefore, by using the DNA marker of the present invention, early diagnosis and screening of individuals with increased muscle fiber type I can easily select high-quality meat with improved meat quality.

According to another aspect of the present invention, the present invention provides a primer for searching for SNPs related to muscle fiber type I increase in muscle of the porcine PPARGC1A gene, consisting of oligonucleotides represented by SEQ ID NO: 2 and SEQ ID NO: 3. In the embodiment of the present invention, a primer for searching for SNP present in the PPARGC1A gene of the present invention was prepared based on the gene sequence registered in Genebank (accession no. AY484500). The primers of the present invention are for amplifying gene fragments containing SNPs, which are not known in the prior art, and enable the preparation of gene fragments containing SNPs at specific sites (SEQ ID NOs: 1 to 678th base) in the porcine PPARGC1A gene sequence. .

According to another aspect of the present invention, the present invention provides a kit for SNP analysis related to muscle fiber type I increase in pigs comprising Alu I, a primer and a PCR reaction mixture and a restriction enzyme for RFLP analysis. The SNP analysis kit of the present invention uses a PCR-RFLP technique, and includes a specific restriction enzyme Alu I to analyze various expressions of genes involved in the regulation of expression of PPARGC1A gene, that is, the muscle fiber composition. Such specific restriction enzymes have been selected by the inventors to identify SNPs that cause changes in muscle fiber composition. Therefore, the kit for searching the SNP of the present invention must include the above components.

In the embodiment of the present invention, genotypes were distinguished by grasping a band pattern appearing differently according to the treatment of Alu I, a restriction enzyme capable of recognizing the SNP site of the present invention. For example, when amplified by primers of SEQ ID NO: 2 and SEQ ID NO: 3, the genotypes are AA, AT, TT, A allele is 61 bp, 60 bp, 31 bp, 27 bp, 21 bp, T allele is 121 It was confirmed that the band appeared at bp, 31 bp, 27 bp, 21 bp (see FIG. 1).

According to another aspect of the present invention, the present invention provides a method for determining whether intramuscular muscle fiber type I increase in pigs comprises the following steps:

a) separating genomic DNA from pigs;

b) amplifying a specific site including the 678th base of the PPARGC1A gene exon 8 represented by SEQ ID NO: 1 using the DNA of step a) as a template;

c) treating the DNA amplified in step b) with restriction enzyme Alu I; And

d) comparing the restriction fragment length polymorphism (RFLP) in the gene fragments obtained in step c).

In the method of the present invention, characterized in that the amplification using the primers shown in SEQ ID NO: 2 and SEQ ID NO: 3 in step b). The DNA amplification of step b) may be any primer capable of amplifying a specific region including SNP, which is the 678th base of the PPARGC1A gene exon 8, but is preferably composed of oligonucleotides represented by SEQ ID NO: 2 and SEQ ID NO: 3 Amplification can be accomplished using primers.

In the method of the present invention, the polymorphism of the gene fragments can be treated with any restriction enzyme capable of recognizing the SNP site, but it is preferable to treat with the restriction enzyme Alu I. The present invention has developed a search method using the PCR-RFLP method as one of the most efficient methods for detecting and diagnosing a marker gene as a single PCR reaction and enzyme treatment in SNP site recognition. Specifically, genotypes were distinguished by grasping band patterns that appear differently according to the treatment of Alu I, a restriction enzyme capable of recognizing the SNP site of the present invention.

Hereinafter, the present invention will be described in more detail with reference to Examples. These embodiments are only for illustrating the present invention, and thus the scope of the present invention is not construed as being limited by these embodiments.

Example 1 Pig Breeding and Genotyping

The pigs used in the experiments were a total of 152 Yorkshire purebred pigs (castration male 54 and female 98) as feed pigs and were fed with the same feed in different cages on the same farm. Three times at 172.2 ± 6.22, 174.6 ± 5.15, and 176.6 ± 9.01 days of age, respectively, they were slaughtered in industrial slaughterhouses under the Korean grading control.

Genomic DNA was collected from muscle samples of the longissimus dorsi using a DNA isolation kit (G-DexTMII, Intronbio., Korea), and the Cys430Ser polymorphism in exon 8 of the pig PPARGC1A gene was Kunej. Genotyping was performed by the PCR-RFLP method of et al (2005).

The primers used in the PCR reaction were forward primer (5'-TAAAGATGCCGCCTCTGACT-3 ') (SEQ ID NO: 2) and reverse primer (5'-) based on the sequence registered in GenBank (Accession no. AY484500) (SEQ ID NO: 1), respectively. CTGCTTCGTCGTCAAAAACA-3 ') (SEQ ID NO: 3) and a total volume of 20 μl 1 μl was added for PCR reaction, and the other reaction additives were 100 ng of genomic DNA, dNTP mix (0.25 mM each), 10 × PCR buffer (300 mM tris-HCl, 300 mM Salts consisting of K ⁺ and NH _4). , 20 mM Mg ²⁺ ), and DNA polymerase 1.25 U (i-MaxTM II, Intronbio., Korea).

For the PCR reaction, a Mastercycler gradient (Eppendorf Co., Germany) instrument was used, and PCR reaction conditions were pre-denaturated at 94 ° C. for 10 minutes; 1 cycle of denaturation at 94 ° C. for 1 minute, annealing at 61 ° C. for 1 minute, and extension of 40 seconds at 72 ° C. for 1 cycle; Finally, the PCR reaction was terminated at 72 ° C. for 10 minutes.

A fragment of the PPARGC1A gene thus obtained (FIG. 2) was treated with an Alu I restriction enzyme (restriction endonuclease) found using a restriction mapper software program (http://www.restrictionmapper.org/).

Alu I restriction enzyme treatment additives include Alu I enzyme 2 μl (2 unit / μl; New England Biolabs, USA), 10 x Enzyme buffer 1.5 μl, 10 x BSA 1.5 μl, 10 x S-adenosylmethionine (SAM, 200 μm / μl ) The total volume was 15 μl with 1.5 μl, ddH ₂ O 5.5 μl and PCR product 3 μl. The mixture was reacted in a 25 ° C. incubator for about 16 hours, and its band pattern was confirmed using poly-acrylamide gel electrophoresis (PAGE) (FIG. 1).

5 microliters of the PCR reaction mixture to 2.5 U restriction enzyme Alu I 5 μl of reaction mixture containing (New England Biolabs, USA) was added. Digestion was performed overnight at 37 ° C., and DNA fragments were separated at 100 V for 30 min in 1 × TAE buffer on 2% agarose gel to confirm the band pattern (see FIG. 1). The genotype of the restriction site is indicated by the T allele for the undigested allele and A allele for the digested allele.

1 shows a PCR-RFLP fragment for an Alu I site located at exon 8 of the PPARGC1A gene. As a size marker, 1kb plus ladder size marker (Invitrogen, USA) was used. The 200 bp PCR product (SEQ ID NO: 4) produced 121 bp, 31 bp, 27 bp, and 21 bp fragments (TT genotype) by restriction enzyme Alu I. The fragments of 31 bp, 27 bp and 21 bp appear identical in all genotypes, and in the AA genotype, 121 bp are cleaved to produce 60 bp and 61 bp fragments. In the case of heterozygotes (AT), both 121 bp, 60 bp, and 61 bp fragments appear. Due to the gel loading characteristics, two segments of 60 bp and 61 bp appear as one band, and segments of 31 bp, 27 bp and 21 bp also appear as one band.

For genotyping results of a total of 152 yolkshire varieties, alleles and genotype frequencies at the Cys430Ser locus of the pig PPARGC1A gene are shown in Table 1 below. The genotype frequency was highest in individuals with AT genotype, and alleles were similarly distributed between A and T allele.

TABLE 1 Genotype and Allele Frequency for Cys430Ser Polymorphism of PPARGC1A Gene in Yorkshire Varieties

N, number of pigs.

Example 2. Analysis of association between genotype, muscle fiber and meat quality

1) muscle fiber characteristics analysis

For muscle fiber characterization, 45 minutes post mortem muscle samples of the dorsum muscles of the thoracic vertebrae were collected, cut into pieces of 0.5- × 0.5- × 1.0-cm, quickly frozen with liquid nitrogen, and -80 ° C until analysis. Were kept at. The transverse section (10 μm thick) was prepared on a cryostat (CM1850, Leica, Germany) machine at 20 ° C. and preincubated with type I, IIa and IIb fibers for myofiblot identification analysis (acid preincubation, pH 4.35). ) Was stained and classified using the actomyosin ATPase method. All samples were characterized by muscle fiber characterization using a CCD camera (charge-coupled device color camera; IK-642K, Toshiba, Japan) and an image analysis system (Image-Pro Plus, Media Cybernetics, USA). . Muscle fiber density (Fiber number per unit area in Table 2) was calculated as the average number of muscle fibers per mm ² per fiber. Total fibers mumbers and fiber density were estimated by multiplying the loin eye area (cm ² ). The results are shown in Table 2 below.

2) meat analysis

Muscle pH was measured at spine 13/14 at 45 minutes post mortem using a spear-type electrode. Lightness was measured at 45 minutes post mortem muscle of the sirloin using a colorimeter (CR-300, Minolta Camera Co., Japan). Drip loss was analyzed by measuring the difference in the amount of muscle sample on the surface in a plastic bag for 48 hours at 2 ° C (Honikel 1987). The results are shown in Table 2 below.

3) Analysis of association between genotype and trait

The general linear analysis model for the association between genotype and trait was as follows. Model: y _ijklm = μ + G _i + S _j + P _k + e _ijkl, where y _ijklm represented the observation, μ the population mean, G _i is a fixed effect, S _j is a fixed effect of sex j according to the genotype i , P _k is the fixed effect of slaughter time k, and e _ijkl is random error. Significant differences were divided by standard errors of mean values, and least squares means were used. Algebraic and angular transformations were applied in the correlation analysis due to composition differences of more than 70%. The statistical program was used as SAS 9.13 (SAS Institute Inc., 2001).

The results of the correlation analysis between genotype and measurement traits such as muscle fiber quality and meat quality are shown in Table 2 below. Significant differences between genotypes in type I number (P <0.01) and area (P <0.05) of the myofibrillar composition showed the function related to type I formation of PPARGC1A. Significant differences (P <0.05) were observed in the size (CSA of fibers, cross-sectional area of fibers) and fiber number per units, suggesting that the candidate gene of the present invention was deeply related to muscle fiber properties. Relevant.

TABLE 2 Effect of Cys430Ser Polymorphism on Muscle Fiber Quality and Meat Quality in Yorkshire Pigs

^{a, b} Least-square means and their standard errors within a row with no common superscript significantly differ.

Levels of significance: NS = not significance, † P <0.1, * P <0.05, ** P <0.01, *** P <0.001.

¹ N showed in bracket is the number of pigs genotyped.

² Cross-sectional area.

In the results of Table 2, there was no difference in the total fibers number in each individual according to the genotype, and the CSA of fibers representing the size of fibers per muscle fiber was large in the TT genotype and the number of fibers per unit area ( Fiber number per unit area was also significant in the TT genotype. The fiber number composition showed a high muscle fiber type I in the AT and TT genotypes with T allele and a high fiber type I type in the AT and TT genotypes. This means that the AT, TT genotype is an increased individual of type I with respect to muscle fiber composition. In terms of meat quality traits, the AT and TT genotypes could be judged as individuals with improved meat quality by showing high pH and lightness, the most important characteristics of meat quality. Therefore, the Cys430Ser polymorphism of the porcine PPARGC1A gene of the present invention, that is, when the A base is substituted with the T base in the 678th sequence of SEQ ID NO: 1 can be judged that muscle fiber type I is increased and the meat quality is improved. Has been shown to be able to be used as a DNA marker to check for increased muscle fiber type I.

In the above, Cys430Ser polymorphism was closely related to meat quality in the composition of type I muscle fibers. It was confirmed that the genotype showed significant difference in pH (P <0.001) and flesh color (P <0.01) of the ventricular myocardium after 45 minutes. Type I had a lower pH and higher brightness than the larger AT and TT genotyping groups. Similar to the results of the present invention, previous studies have reported that the increase in the number and size composition of type I muscle fibers is associated with a 45-minute increase in mesenchymal pH and reduction in color (Ryu & Kim 2005; Ryu et al. 2008). In addition, abnormal pork such as PSE has been reported to have lower pH, higher brightness, and higher post-mortem metabolic rate than normal pork (Ryu & Kim 2006). Therefore, the AT, TT genotypes of the type I increased individuals according to the present invention can be determined to exhibit improved meat quality.

As described above, according to the present invention, the Cys430Ser mutation of exon 8 of the PPARGC1A gene can be judged to affect the type I muscle fiber formation of porcine longissimus dorsi muscles. It can be inferred that the change in affects the post-metabolism of muscles and affects the final meat quality. Therefore, the genetic variation (SNP) of the present invention has a function related to the muscle fiber composition of pigs, and is expected to be useful as a DNA marker related to meat improvement through this property.

[references]

Berchtold, MW, H. Brinkmeier, et al. (2000). "Calcium Ion in Skeletal Muscle: Its Crucial Role for Muscle Function, Plasticity, and Disease." Physiol. Rev. 80 (3): 1215-1265.

Brocks, L., RE Klont, et al. (2000). "The effects of selection of pigs on growth rate vs leanness on histochemical characteristics of different muscles." J. Anim Sci. 78 (5): 1247-1254.

Cameron, N., P. Warris, et al. (1990). "Comparison of Duroc and British Landrace pigs for meat and eating quality." Meat Sci 27: 227-247.

Erkens, T., M. Van Poucke, et al. (2006). "Development of a new set of reference genes for normalization of real-time RT-PCR data of porcine backfat and longissimus dorsi muscle, and evaluation with PPARGC1A." BMC Biotechnol 6: 41.

Honikel, KO (1987). How to measure the water-holding capacity of meat Recommendation of standardized methods. Evaluation and control of meat quality in pigs . Dordrecht, The Netherlands: Martinus Nijhoff, In PV Tarrant, G. Eikelenboom, & G. Monin (Eds.): 129-142.

Jacobs, K., G. Rohrer, et al. (2006). "Porcine PPARGC1A (peroxisome proliferative activated receptor gamma coactivator 1A): coding sequence, genomic organization, polymorphisms and mapping." Cytogenet Genome Res 112 (1-2): 106-113.

Karlsson, A., AC Enfalt, et al. (1993). "Muscle histochemical and biochemical properties in relation to meat quality during selection for increased lean tissue growth rate in pigs." J. Anim Sci. 71 (4): 930-938.

Klont, RE, L. Brocks, et al. (1998). "Muscle fiber type and meat quality." Meat Science 49 (Supplement 1): S219-S229.

Kunej, T., XL Wu, et al. (2005). "Frequency distribution of a Cys430Ser polymorphism in peroxisome proliferator-activated receptor-gamma coactivator-1 (PPARGC1) gene sequence in Chinese and Western pig breeds." J Anim Breed Genet 122 (1): 7-11.

Larzul, C., L. Lefaucheur, et al. (1997). "Phenotypic and genetic parameters for longissimus muscle fiber characteristics in relation to growth, carcass, and meat quality traits in large white pigs." J. Anim Sci. 75 (12): 3126-3137.

Lin, J., H. Wu, et al. (2002). "Transcriptional co-activator PGC-1 [alpha] drives the formation of slow-twitch muscle fibers." Nature 418 (6899): 797-801.

Puigserver, P., Z. Wu, et al. (1998). "A Cold-Inducible Coactivator of Nuclear Receptors Linked to Adaptive Thermogenesis." Cell 92 (6): 829-839.

Rehfeldt, C. and I. Fiedler (1984). "[Postnatal development of muscle fibers in growing skeletal muscle of laboratory mice]." Arch. Exp. Vet. med. 38: 178-192.

Rehfeldt, C., I. Fiedler, et al. (2000). "Myogenesis and postnatal skeletal muscle cell growth as influenced by selection." Livestock Production Science 66 (2): 177-188.

Ryu, YC, YM Choi, et al. (2008). "Comparing the histochemical characteristics and meat quality traits of different pig breeds." Meat Sci. doi: 10.1016 / j.meatsci.2007.12.020.

Ryu, YC and BC Kim (2005). "The relationship between muscle fiber characteristics, postmortem metabolic rate, and meat quality of pig longissimus dorsi muscle." Meat Science 71 (2): 351-357.

Ryu, YC and BC Kim (2006). "Comparison of histochemical characteristics in various pork groups categorized by postmortem metabolic rate and pork quality." J. Anim Sci. 84 (4): 894-901.

Stachowiak, M., M. Szydlowski, et al. (2006). "SNPs in the Porcine PPARGC1a Gene: Interbreed Differences and Their Phenotypic Effects." Cell Mol Biol Lett .

te Pas, MF, FJ Verburg, et al. (2000). "Messenger ribonucleic acid expression of the MyoD gene family in muscle tissue at slaughter in relation to selection for porcine growth rate." J. Anim Sci. 78 (1): 69-77.

1 shows a PCR-RFLP fragment for an Alu I site located at exon 8 of the PPARGC1A gene. As a size marker, 1kb plus ladder size marker (Invitrogen, USA) was used.

2 shows that the 678th base in the base sequence of the PPARGC1A gene exon 8 is SNP substituted with A / T.

<110> Korea University Industrial and Academic Collaboration Foundation <120> DNA markers for detecting increase of muscle fiber type I within          porcine muscle <160> 4 <170> KopatentIn 1.71 <210> 1 <211> 2098 <212> DNA <213> Sus scrofa <400> 1 gggcgtccga agtacaaaat atcatcactg cgactgmatc ctcccgtcgt aagatcccgt 60 cttttcttgg ggtcaggggc ttcgggcaca gagtcacttt ctcaggtctt gtcatgaatt 120 tagcttcctc ctttttcaca gcgttttaac cactgtccgc aatctcagaa acgttaagaa 180 aacgtaccct ggcatttacc attctccctg tctcttctct ctcccccccc tacccttttt 240 ttctcccccc tcttatttta ggcctaactc cacccacaac tcctcctcat aaagccaacc 300 aagataaccc ttttagggct tctccaaagc tgaagccccc ttgcaagact gtggtacctc 360 cgccatcgaa gaagacccgg tacagtgagt cttcggggac ccacggcaac aactccacca 420 agaaagggcc cgagcagtcc gagctgtacg cgcagctcag caagacgtcc gcgctcggcg 480 gcggacacga ggaacggaag gccaggcggc ccagtctgcg gctatttggt gaccatgact 540 attgtcagtc gattaattcc aaagcggaaa tcctcatcaa tatatcgcag gagctccacg 600 actccagaca actagactct aaagatgccg cctctgactg gcagaggcag atgtgttctt 660 ccacagactc agaccagtgc tacctgaccg agacgtcgga ggcgagcagg caggtctctc 720 cgggcagcgc ccgaaaacag ctccaagacc aggaaatccg agccgagctg aacaagcact 780 tcggtcatcc cagtcaagct gtttttgacg acgaagcaga caagaccagt gaactgaggg 840 acagtgattt cagtaacgaa caattctcca aactacctat gtttataaat tcaggactag 900 ccatggatgg cctgtttgat gacagcgaag atgaaagtga taaactgaac tccccttggg 960 atggcacgca gtcctattca ttgttcgatg tgtcgccttc ttgttcttct tttaactctc 1020 cgtgtagaga ttccgtatca ccacccaaat ccttattttc tcaaagaccc caaaggatgc 1080 gctctcgttc aaggtccttt tctcaacaca ggtcgtgttc tcgatcacca tattccaggt 1140 caagatcaag gtccccaggc agtagatcct cttcaaggta aaccttggca aaacctaccc 1200 aaagcctaag gcatccccag gaagtcaaaa aaaaaaaaaa ttcaaaaccc acaggcatcc 1260 ggtgctgcca gaactatgag attgttggtg cccatacatt tctccccttt gtgtcttaaa 1320 tctgtcgtcc ttgggcacag ggccttgagt cagtcagatc agagggagca atttgtaaga 1380 aacctgccca tctagctttt cccattacct gcagagctgc cttagtacca ggatagttgc 1440 gatccatgcg agataaagct gacagcatcc cagactggcg tttgcgggag ataccgcgag 1500 agaacgcatc cctcccagcc ctctaaccaa ctagctgatc acagacggtc actgggaaac 1560 ctcactagtt actgtgaagc aaaagcctgt ttatgtgata cggagctcca ggggccattc 1620 agtcatgccc atggtaatct ggggttcacg ggagttgcaa tgccctttgt gtgtgtcccc 1680 ctctcttgta gatcttgcta ctactctgag tcaggccact gcagacaccg cacgcaccga 1740 aattctcccc tgtgcgccag atcacgttca agatctccct acagccggcg gcccaggtaa 1800 tgaccttcca ctccttgcat ccccaccctt ggatgccgag tcgtggcccc aagccctgcc 1860 ctctccccag tttgtgattc attctgcggg tgcagatgga ggtggtggac tgaatgctga 1920 agttctgttt ctcaggtatg acagctacga ggaatatcag cacgagaggc tgaagaggga 1980 agaataccgc agagagtatg agaagcggga gtctgaaagg gccaagcaga gggagaggca 2040 gaggcagaag gcaattgtaa gcagcctgga gctcactttc gttttacagg agggtttc 2098 <210> 2 <211> 20 <212> DNA <213> Artificial Sequence <220> <223> Forward primer for PCR-RFLP <400> 2 taaagatgcc gcctctgact 20 <210> 3 <211> 20 <212> DNA <213> Artificial Sequence <220> <223> Reverse primer for PCR-RFLP <400> 3 ctgcttcgtc gtcaaaaaca 20 <210> 4 <211> 200 <212> DNA <213> Sus scrofa <400> 4 taaagatgcc gcctctgact ggcagaggca gatgtgttct tccacagact cagaccagtg 60 ctacctgacc gagacgtcgg aggcgagcag gcaggtctct ccgggcagcg cccgaaaaca 120 gctccaagac caggaaatcc gagccgagct gaacaagcac ttcggtcatc ccagtcaagc 180 tgtttttgac gacgaagcag 200  

Claims (7)

In the nucleotide sequence of SEQ ID NO: 1, consisting of 15 to 500 consecutive sequences including the 678th T base, DNA marker for confirming whether muscle fiber type I increase in pigs. The method of claim 1, wherein the DNA marker is a DNA marker, characterized in that to improve the meat quality by increasing the ratio of muscle fiber type I in the muscle fiber composition of the pig. 3. The DNA marker according to claim 2, wherein the improvement of meat quality comprises an increase in pH and a decrease in meat color. A primer for searching for an SNP related to muscle fiber type I increase in the porcine PPARGC1A gene, consisting of oligonucleotides represented by SEQ ID NO: 2 and SEQ ID NO: 3. SNP analysis kit related to muscle fiber type I increase in pigs comprising the primer of claim 4 and a PCR reaction mixture and Alu I, a restriction enzyme for RFLP analysis. A method for determining whether muscle fibers type I increase in pigs, comprising the following steps: a) separating genomic DNA from pigs; b) amplifying a specific site including the 678th base of the PPARGC1A gene exon 8 represented by SEQ ID NO: 1 using the DNA of step a) as a template; c) treating the DNA amplified in step b) with restriction enzyme Alu I; And d) comparing the restriction fragment length polymorphism (RFLP) in the gene fragments obtained in step c). The method of claim 6, wherein the amplification using the primers represented by SEQ ID NO: 2 and SEQ ID NO: 3 in step b).

Images