KR20160049177A - Prediction method for swine fecundity using gene expression profile - Google Patents

Abstract

The present invention relates to a method for estimating fecundity in swine by applying differentially expressed gene (DEG). In order to improve, as genetic resources, varieties of black pigs showing excellent fecundity, an analysis is carried out on RNA derived from the placenta of both less fecund mother pigs and highly fecund mother pigs so as to obtain the DEG, and a correlation between the gene and the fecundity is figured out, making it possible to provide a diagnostic technology for setting up lineage for black pigs showing excellent fecundity.

Description

[0001] Description [0002] Prediction method for swine fecundity using gene expression profile [

The present invention relates to a method for predicting the number of pigs by using the expression profile of a gene, and more particularly, to a composition for predicting the number of pigs by using a gene expression profile and a method for predicting the number of pigs.

The number of pigs is estimated to be very high compared to other traits, but relatively low heritability and technical limitations make it difficult to improve. In recent years, the importance of pork production capacity testing project to improve the number of living quarters and the number of reasons has been recognized. Therefore, in Korea as well as in Europe, a group of sows with a large number of siblings is formed, and the group continues to develop superior systems. This is called a hyper-prolific line. The improvement of sows is mainly focused on reproductive ability and sowing robustness. The main items are the number of litter size, the number of start of breastfeeding, birth weight, 21 day old body weight, 21st birthday number. In the developed countries such as USA, UK and Japan, many studies have been actively carried out on the improvement of the number of pigs by importing the genes of Meishan species, a Chinese native species, in order to cultivate multi-acid strains. However, have.

Numerous cited documents and patent documents are referred to and cited throughout this specification. The disclosures of the cited documents and patents are incorporated herein by reference in their entirety to more clearly describe the state of the art to which the present invention pertains and the content of the present invention.

Korean Registered Patent No. 0444160 (2004.08.02)

Under these technical backgrounds, the present inventors have made intensive efforts to accomplish the present invention. DISCLOSURE OF THE INVENTION The object of the present invention is to obtain DEG (Differentially Expressed Gene) by analyzing RNA from placenta of sows having a high number of siblings and sows with an inferior number of sows, And to provide a diagnostic technology for the composition of the black pork system having a high number of westerners.

Other objects and advantages of the present invention will become more apparent from the following detailed description of the invention, claims and drawings.

According to one aspect of the present invention there is provided a method of screening for the presence of pig genes CHI3L1, EGR2, HBA, LIPG, NFKBIZ, PDPN, PHEROC, PLAT, PLAUR, PPP1R15A, SPRY2, XIRP1, ZNF385D, A2ML1, ACTG2, ADAMTS8, ALDH1A2, ANKH, ANKRD9, APOB, ATF3, BCAR1, BCAS1, CD86, CDH17, CDHR2, CDHR5, CHPF, CHRDL1, CHST3, CNN1, COL1A2, COL3A1, COL4A2, COL5A1, COL7A1, CREB3L3, CRNN, CYP2W1, DAGLA, DAO, DSG1, EMILIN1, EPS8L3, IGFBP5, IGSF23, IL21R, ITIH4, KCNK5, KRT1, KRT13, KRT14, KRT15, KRT20, KRT4, KRT5, KRT77, LCN15, GFG1, GFG1, RAVP2, RBP4, REG4, RAVP2, RBP4, RBP2, RBP4, RBP4, RBP2, MUC13, MUC4, MUC4, MUC4, MUC4, MUC4, RLCK4, ROS1, SCARA5, SDK1, SH3GL2, SLC15A1, SLC16A3, SLC2A8, SLC38A10, SLC45A3, SLC45A4, SLC5A1, SLC5A6, SLC6A19, SLIT3, SPOCK3, SYNM, THBS2, TM4SF20, TOR4A, TRIM3, UPK1A, USH1C, VAT1L, ZFYVE21 < / RTI > < RTI ID = 0.0 > A composition for predicting the number of live pigs of pigs may be provided, including a preparation for measuring the present level.

According to another aspect of the present invention, there is provided a kit for estimating the number of pigs of a pig comprising the composition.

In one embodiment, the kit may be an RT-PCR kit, a microarray chip kit, or a protein chip kit.

According to another aspect of the present invention, there is provided a method for quantifying the expression level of each gene, comprising: extracting mRNA from two or more pigs and pooling the mRNA; And

Wherein at least one gene of CHI3L1, EGR2, HBA, LIPG, NFKBIZ, PDPN, PHEROC, PLAT, PLAUR, PPP1R15A, SPRY2, XIRP1 and ZNF385D is expressed higher than the average expression amount in each quantified gene, CYP2W1, CYP2W1, CYP2W1, CYP2W1, CYP2W1, CYP2W1, CYP2W1, CYP2W1, CYP2W1, DAGLA, DAO, DSG1, EMILIN1, EPS8L3, EVPL, F10, FAM174B, FAM89A, FBLN5, FBP2, FHL1C, GJB1, GLDN, GPR153, HAL, HNF4A, IGFBP5, IGSF23, IL21R, ITIH4, KCNK5, KRT1, KRT13, KRT14, MCPD1, MMP8, MPV17L2, MUC13A, MUC4, MXRA5, NPC1L1, PAPSS2, PCDH1, PCK1, PIM3, PPL, PPP1R3D, MKD15, KRT20, KRT4, KRT5, KRT77, LCN15, LGALS2, LOX, LREAP1, LTBP2, MAMDC4, MAOB, PRP1, PRKCDBP, PRKG2, RBP2, RBP4, REG4, RIPK4, ROS1, SCARA5, SDK1, SH3GL2, SLC15A1, SLC16A3, SLC2A8, SLC38A10, SLC45A3, SLC45A4, SLC5A1, SLC5A6, SLC6A19, SLIT3, SPOCK3, SYNM, THBS2, TM4SF20, TOR4A, TRIM3, UPK1A, USH1C, VAT1L, VIL 1 < / RTI > and ZFYVE21 is expressed less than the average expression amount, to a pig having a higher number of pigs.

According to the present invention, it is possible to effectively predict the number of hatchlings of pigs, and to form a pig system having an excellent number of hatchlings.

Figure 1 shows the results of the DEG analysis for the high population numbers (TN1308R4083) and low population numbers (TN1307R3720) groups with various results.
FIG. 2 shows the genes and genes of genes having significance associated with the molecular function in DEG.
FIG. 3 shows genes and genes of genes having significance associated with the biological process in DEG.
FIG. 4 shows genes and genes of genes having significance associated with cellular components in DEG.

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

According to one aspect of the present invention there is provided a method of screening for the presence of pig genes CHI3L1, EGR2, HBA, LIPG, NFKBIZ, PDPN, PHEROC, PLAT, PLAUR, PPP1R15A, SPRY2, XIRP1, ZNF385D, A2ML1, ACTG2, ADAMTS8, ALDH1A2, ANKH, ANKRD9, APOB, ATF3, BCAR1, BCAS1, CD86, CDH17, CDHR2, CDHR5, CHPF, CHRDL1, CHST3, CNN1, COL1A2, COL3A1, COL4A2, COL5A1, COL7A1, CREB3L3, CRNN, CYP2W1, DAGLA, DAO, DSG1, EMILIN1, EPS8L3, IGFBP5, IGSF23, IL21R, ITIH4, KCNK5, KRT1, KRT13, KRT14, KRT15, KRT20, KRT4, KRT5, KRT77, LCN15, GFG1, GFG1, RAVP2, RBP4, REG4, RAVP2, RBP4, RBP2, RBP4, RBP4, RBP2, MUC13, MUC4, MUC4, MUC4, MUC4, MUC4, RLCK4, ROS1, SCARA5, SDK1, SH3GL2, SLC15A1, SLC16A3, SLC2A8, SLC38A10, SLC45A3, SLC45A4, SLC5A1, SLC5A6, SLC6A19, SLIT3, SPOCK3, SYNM, THBS2, TM4SF20, TOR4A, TRIM3, UPK1A, USH1C, VAT1L, ZFYVE21 < / RTI > < RTI ID = 0.0 > A composition for predicting the number of live pigs of pigs may be provided, including a preparation for measuring the present level.

Here, measuring the above expression level may be a measure of the level of mRNA or protein.

Measuring the level of mRNA hereinabove can be analyzed by any of the known methods including RT-PCR, competitive RT-PCR, real-time RT-PCR, RNase protection assay, Northern blotting, DNA microarray and the like. Preferably, mRNA isolated from the biological sample or cDNA derived therefrom is hybridized on a microarray in which a probe specific to one or more marker genes selected from the group consisting of the genes is immobilized, and the resulting degree of hybridization is measured . The hybridization degree can be measured by any measurement method known in the art such as fluorescence measurement and electrical measurement. In this case, the probe or the target nucleic acid may be labeled with a detectable appropriate label. Here, the cDNA may be directly amplified by RT-PCR using a pair of sense and antisense primers targeting at least one marker gene selected from the group consisting of the genes as primers.

In the present invention, the above-mentioned "measurement of the level of the protein" may be performed using any of known protein measurement or detection methods. For example, an assay method using an antibody that specifically binds to a protein expressed from one or more marker genes selected from the group consisting of the genes may be used. Methods for analyzing proteins using antibodies include Western blotting, ELISA, radioimmunoassay, radial immunodiffusion, Oucheronin immunodiffusion, rocket immunoelectrophoresis, tissue immunostaining, immunoprecipitation assays, complement fixation assays, FACS But are not limited to these examples. Such ELISAs include direct ELISA, indirect ELISA, direct sandwich ELISA, indirect sandwich ELISA, and the like. Western blotting refers to separation of whole proteins, electrophoresis, separation of proteins according to their size, transfer to a nitrocellulose membrane, reaction with the antibody, and quantification of the amount of the produced antigen-antibody complex by using labeled antibodies It is a way to confirm. In addition, methods for measuring protein levels include methods using enzymes, substrates, coenzymes, and ligands that specifically bind to target proteins.

In the present invention, the expression level of the gene is determined by measuring the amount of the amplification product obtained by nucleic acid amplification performed by RT-PCR, using RNA isolated from the sample as a template Lt; / RTI >

The composition may further include a reagent necessary for hybridization with the marker gene in the sample or the nucleic acid expression product expressed therefrom. In addition, the composition may further comprise a buffer, a solvent, etc., which stabilizes the probe and becomes a reaction medium.

Throughout this specification, the term "probe" is an oligonucleotide that is capable of binding to a target nucleic acid in a base-specific manner, as a nucleic acid strand partially or completely complementary to the target nucleic acid. Preferably, it is an oligonucleotide that is completely complementary to the target nucleic acid. The probe includes not only nucleic acid but also any nucleic acid derivative known in the art which is capable of complementary binding including a peptide nucleic acid.

The binding of the probe to the target nucleic acid (generally, also referred to as hybridization) occurs in a sequence-dependent manner and can be performed under various conditions. Generally, the hybridization reaction occurs at a temperature about 5 ° C below the Tm for a particular sequence at a specific ionic strength and pH. The Tm means that 50% of the probe complementary to the target sequence is bound to the target sequence. An example of the hybridization reaction conditions may be a pH 7.0 to 8.3, 0.01 to 1.0 M Na + ion concentration. In addition, in order to enhance the specificity of the target nucleic acid and the probe, it is necessary to carry out the hybridization under the condition that the binding of the probe nucleic acid and the target nucleic acid becomes unstable, for example, in the presence of a high temperature, high concentration of a destabilizer (for example, formamide) .

The length of the probe is capable of specifically binding to the target nucleic acid sequence, and includes polynucleotides of any length. For example, the length of the probe may be a length of 7 to 200 nucleotides, 7 to 150 nucleotides, 7 to 100 nucleotides, 7 to 50 nucleotides, or a single strand of a full-length gene, but is not limited thereto.

The probe may be labeled with a detectable label. The detectable label includes a fluorescent label such as Cy3 or Cy5, a radioactive label, an enzyme for converting the substrate into a coloring material, and the like, but the present invention is not limited to these examples.

According to another aspect of the present invention, there is provided a kit for estimating the number of pigs of a pig comprising the composition.

In one embodiment, the kit may be an RT-PCR kit, a microarray chip kit, or a protein chip kit.

According to another aspect of the present invention, there is provided a method for quantifying the expression level of each gene, comprising: extracting mRNA from two or more pigs and pooling the mRNA; Wherein at least one gene of CHI3L1, EGR2, HBA, LIPG, NFKBIZ, PDPN, PHEROC, PLAT, PLAUR, PPP1R15A, SPRY2, XIRP1 and ZNF385D is expressed in a higher level than the average expression amount in each quantified gene, , ACTG2, ADAMTS8, ALDH1A2, ANKH, ANKRD9, APOB, ATF3, BCAR1, BCAS1, CD86, CDH17, CDHR2, CDHR5, CHPF, CHRDL1, CHST3, CNN1, COL1A2, COL3A1, COL4A2, COL5A1, COL7A1, CREB3L3, CRNN, CYP2W1 , DAGLA, DAO, DSG1, EMILIN1, EPS8L3, EVPL, F10, FAM174B, FAM89A, FBLN5, FBP2, FHL1C, GJB1, GLDN, GPR153, HAL, HNF4A, IGFBP5, IGSF23, IL21R, ITIH4, KCNK5, KRT1, KRT13, KRT14 , KLT15, KRT20, KRT4, KRT5, KRT77, LCN15, LGALS2, LOX, LREAP1, LTBP2, MAMDC4, MAOB, MFSD10, MMP8, MPV17L2, MUC13A, MUC4, MXRA5, NPC1L1, PAPSS2, PCDH1, PCK1, PIM3, PPL, PPP1R3D , PRP1, PRKCDBP, PRKG2, RBP2, RBP4, REG4, RIPK4, ROS1, SCARA5, SDK1, SH3GL2, SLC15A1, SLC16A3, SLC2A8, SLC38A10, SLC45A3, SLC45A4, SLC5A1, SLC5A6, SLC6A19, SLIT3, SPOCK3, SYNM, THBS2, TM4SF20 , TOR4A, TRIM3, UPK1A, USH1C, VAT1L, A step of predicting the number of pigs of a pig, which comprises predicting a case where at least one gene of VIL1 and ZFYVE21 is expressed in an amount less than the average expression amount, into a pig having a higher number of pigs.

The diagnosis of the present invention can be performed as follows.

The present invention obtains a gene profile of a pig that does not know the number of hatchlings, and the average expression level of genes obtained from a plurality of pigs is used as a control group. The population pig for calculating the average expression amount is preferably the same species as it is desirable to have a similar genome, and it is preferable that the pig is collected from different households in order to ensure diversity of gene expression among individuals. It is also desirable to obtain an average expression amount from as many individuals as possible in order to increase the accuracy and significance of the statistics.

Expression of 'above the average expression level' above means that expression of the gene is significantly increased compared with the average expression level of the genes collected from two or more pigs. That is, when a gene profile of pigs to be tested is obtained in which the number of hatchlings is not known, the list of genes having increased expression in pigs of a high number of pigs and the genes whose expression has decreased in the piglets of the above-mentioned genes is referred to, The amount of the pigs to be tested can be predicted by examining whether or not the amount of the pigs is significantly increased.

The gene expression profiles obtained from the specimens of the present invention can be predicted to have a higher or lower number of embryos than the control group.

The prediction may be to statistically analyze and process aspects of the sample gene profile.

At this time, various methods for predicting the number of residents more accurately can be used.

For example, the DEGs listed in the following Tables 1 and 2 are determined according to the order of the differences in the expression levels, and the profiles of the genes obtained from the specimens are examined to determine the degree of importance according to the sequence, It can be applied to the prediction of the number.

The DEGs shown in the following Tables 1 and 2 can be classified into categories according to the gene ontology, the profiles of the genes obtained from the specimens can be examined, and the DEGs can be applied to the prediction of the number of animals by differentiating them according to categories.

Hereinafter, the present invention will be described in more detail with reference to Examples. It should be understood, however, that these examples are for illustrative purposes only and are not to be construed as limiting the scope of the present invention.

Experimental Method

Placenta sampling and mRNA  Sequencing

The placenta was collected from black pork sow immediately after delivery in 12 higher litter size and 7 lower litter size by referring to the Lambs Associated Chart, and a certain amount of the placenta was cut at the same site, Respectively. Total RNA was isolated from the placental tissue using a TRI reagent (Molecular Research Center, Cincinnati, Ohio, USA) after cutting. mRNA was isolated by RNA-Seq sample purification kit (Illumina, Inc., San Diego, Calif., Jung et al., 2012).

RNA purification was determined by analyzing 1 μl of total RNA extract on a NanoDrop 8000 spectrophotometer. The integrity of the total RNA was checked with an Agilent Technologies 2100 Bioanalyzer and the RNA integrity index (RIN) was based on a value of 8 or greater. The mRNA sequencing library was prepared according to the manufacturer's instructions (Illumina TruSeq RNA Prep kit v2). The mRNA was repeatedly purified twice using 2 μg of total RNA and purified and fragmented using a magnetic bead with poly-T oligonucleotide attached thereto. The fragmented RNA fragment was primed with random hexamers and reverse transcribed with 1 ^st cDNA with reverse transcriptase. The RNA template was then removed and dsDNA was synthesized. Enrichment of the purified cDNA was performed by end repair, A-tailing, adapter ligation, cDNA template purification and PCR. The quality of the amplified library was confirmed by capillary electrophoresis (Bioanalyzer, Agilent).

QPCR was performed using SYBR Green PCR Mastermix (Applied Biosystems) and the pool generated was then combined with the tagged library in this same amount.

Cluster generation was performed in a flow cell of the cBot automated cluster generation system (Illumina). Flow cells were loaded into a HISEQ 2500 sequencing system (Illumina) and sequenced with a 2x100 bp lead length.

RNA  Sequencing results

RNA was separated from each of the three strains of low abundance (TN1307R3720) and high abundance (TN1308R4083), pooled and RNA-seq was performed. As a result, the total number of leads was 47,641,439 (low number of inhabitants) and 45,940,962 (high number of inhabitants), and the proper paired read was 29,141,890 (61.17%) and 28,078,596 (61.12% (Fig. 1).

In order to remove the low quality sequence, the lead containing the nucleotide represented by N in the sequence information of 10% or more of the entire sequence or the base of less than 40% of the base of Q20 was removed, and the lead having the average quality of less than Q20 was also removed Respectively. The entire filtering process was performed by an internally generated program. The reference genome used for sequence alignment and analysis was the information provided in Ensembl (Flicek P. et al., 2013) and version 72 was used. The filtered sequence was aligned to the genomic sequence using STAR 2.3.0e (Dobin et al, 2013) and the gene information of ensembl version 72 was used in the sequence alignment. The number of Sus scrofa genes by the reference genome was predicted to be 25,323, and the number of transcripts was 30,587.

DEG  Analysis

The expression level was measured using Cufflinks v2.1.1 (Trapnell C. et al, 2010). In order to measure the expression level, the gene information of the ensembl 72 version was used, and the non-coding gene region was excluded from the expression amount measurement using the -mask option. In order to increase the accuracy of the expression measurement, multi-read-correction and frag-bias-correct options were additionally used, and other options were used as default values.

For specific expression gene analysis, the lead number of each gene was calculated using HTSeq-count v0.5.4p3 (Anders S. et al, 20140), and the intersection-nonempty rule and the pair- End (Paired-end) sequence. Specific gene expression analysis using TCC (Sun J. et al, 2013) was performed using the calculated lead number of each gene. The TCC option used the iDEGES / edgeR method with consideration of repetition, and the specific expression gene selection was set at a reference value of less than 0.05 based on the Q-value corrected for errors caused by multiple-testing.

As a result of DEG analysis, 278 genes were found to be significantly involved in DEG. Compared to the low - abundance group of high - abundance genes, there were 37 high - expression genes and 241 low - expression genes. The results of the clustering according to the DEG are shown in FIG. 1, and it can be seen that clustering is well performed for both groups.

Analysis of the Pearson correlation of DEG showed a high correlation of 0.962 or higher.

As a result of the analysis of the read gene, the total genes detected in both groups were 19,545, among which 14,824 and 14,511 genes were mapped in the low and high expression groups, respectively . No new gene was found.

Gene Ontology Analysis

The Gene Ontology (GO) classifies genes into three categories: biological processes (BP), cell components (CC), and molecular functions (MF) , And provides information about the functions of the currently selected gene. In order to know the characteristics of the genes selected through specific expression gene analysis, a tendency analysis using gene ontology was performed. Using Fisher's accuracy test (Fisher RA, 1922), significant gene ontology classification Respectively. DEG genes were classified by function according to the criteria of molecular functions, biological processes, and cellular components. Of the total 278 DEGs, 117 DEGs were available for gene ontology analysis and 1,727 gene ontologies were available. Of these, the significant DEG categories were 3, 6, and 6, respectively, for molecular function, biological process, and cellular component criteria. The number of genes involved in binding in the molecular function was the largest, 179, and categories were classified as molecular functions and carbon-carbon lyase activity. Specific functional classifications were structural molecule activity, fructose-bisphosphate aldolase activity, and aldehyde-lyase activity. Structural molecule activity was associated with 14 genes, including DES, three of which were uncharacterized. The fructose biphosphate agarase activity and the aldehyde-decomposing activity were found to work with ALDOB.

Biological processes have a high degree of distribution, including single organism processes, cellular processes, metabolic processes, and biological regulation. This category includes single-multicellular organism processes, organic substance metabolic processes, biological processes, cellular response to inorganic substances, chemical stimuli Cellular response to chemical stimulus, establishment of localization, and so on. The associated genes include a total of 13 genes including APOA4 and one uncharacterized state gene. Cellular components appeared in the order of cells, cellular parts, cell membranes, and organelles. The categories were classified as cell parts, keratin filaments, and cellular components. Ten genes, including COL18A1, are involved.

Number of Sanjas  Analysis of expression changes of related genes

Analysis of 278 DEG data was performed based on the results of NCBI data analysis. As a result, up-regulated genes were associated with 13 genes and down-regulated genes were associated with 108 genes.

Among the up-regulated genes, PHEROC (Sus scrofa pheromaxein C subunit) was the highest at 2 ^2.51 (5.7) times and XIRP1 (Sus scrofa xin actin-binding repeat containing 1) was ^2.42 (5.35) times higher.

Among the down-regulated genes, KRT1 (Sus scrofa keratin 1) was the lowest at 2 ^-10.2 (8.5 x 10 ^-4 ) times. The genes showing a value of 2 ^-7 (7.8 x 10-3) or less are HNF4A (Sus scrofa hepatocyte nuclear factor 4 alpha), IGSF23 (Sus scrofa immunoglobulin superfamily, member 23), LGALS2 (Sus scrofa lectin, galactoside-binding , soluble 2), RBP2 (Sus scrofa retinol binding protein 2), and TM4SF20 (Sus scrofa transmembrane 4L six family member 20).

Taken together, the placenta from the group according to the number of living organisms was collected and subjected to RNA sequencing, resulting in a total of 278 DEGs. Of these, total 117 DEGs were analyzed through gene ontology. There were 35 genes with significance for gene ontology. As a result of analyzing the genes related to the number of hatchlings, 13 genes were up-regulated in the high number of hatchings and 108 genes were down-regulated.

No Name Description Value_1 Value_2 log2 p-value One CHI3L1 Sus scrofa chitinase 3-like 1 (cartilage glycoprotein-39) (CHI3L1), mRNA. [Source: RefSeq mRNA; Acc: NM_001001540] 1,460,000 3,500,000 1.26 0.0017 2 EGR2 Sus scrofa early growth response 2 (EGR2), mRNA. [Source: RefSeq mRNA; Acc: NM_001097488] 420,000 1,290,000 1.62 0.0002 3 HBA Hemoglobin subunit alpha [Source: UniProtKB / Swiss-Prot; Acc: P01965] 6,760,000 14,900,000 1.14 0.0033 4 LIPG Sus scrofa lipase, endothelial (LIPG), mRNA. [Source: RefSeq mRNA; Acc: NM_001243029] 380,000 1,080,000 1.51 0.0007 5 NFKBIZ nuclear factor of kappa light polypeptide gene enhancer in B-cells inhibitor, zeta [Source: HGNC Symbol; Acc .: 29805] 125,000 452,000 1.85 0.0005 6 PDPN podoplanin [Source: HGNC Symbol; Acc: 29602] 213,000 623,000 1.55 0.0014 7 PHEROC Sus scrofa pheromaxein C subunit (PHEROC), mRNA. [Source: RefSeq mRNA; Acc: NM_001123161] 18.900 108,000 2.51 0.0055 8 PLAT Sus scrofa plasminogen activator, tissue (PLAT), mRNA. [Source: RefSeq mRNA; Acc: NM_214054] 2,210,000 5,010,000 1.18 0.0029 9 PLAUR plasminogen activator, urokinase receptor [Source: HGNC Symbol; Acc: 9053] 595,000 1,560,000 1.39 0.0011 10 PPP1R15A Uncharacterized protein [Source: UniProtKB / TrEMBL; Acc: F1RIN9] 2,550,000 5,240,000 1.04 0.0082 11 SPRY2 Uncharacterized protein [Source: UniProtKB / TrEMBL; Acc: F1RP67] 251,000 646,000 1.37 0.0042 12 XIRP1 Sus scrofa xin actin-binding repeat containing 1 (XIRP1), mRNA. [Source: RefSeq mRNA; Acc: NM_001143928] 19,900 107,000 2.42 0.0071 13 ZNF385D Uncharacterized protein [Source: UniProtKB / TrEMBL; Acc: F1RS58] 453,000 1,030,000 1.18 0.0075

No Name Description Value_1 Value_2 log2 p-value One A2ML1 Uncharacterized protein [Source: UniProtKB / TrEMBL; Acc: F1SLW8] 159,000 26.100 -2.61 0.0008 2 ACTG2 Uncharacterized protein [Source: UniProtKB / TrEMBL; Acc: F1SLG5] 4,250,000 1,030,000 -2.04 0.0000 3 ADAMTS8 Uncharacterized protein [Source: UniProtKB / TrEMBL; Acc: F1S6D3] 546,000 78.400 -2.8 0.0000 4 ALDH1A2 Uncharacterized protein [Source: UniProtKB / TrEMBL; Acc: I3LK62] 135,000 25.100 -2.43 0.0029 5 ANKH Uncharacterized protein [Source: UniProtKB / TrEMBL; Acc: F1SRN1] 2,270,000 1,090,000 -1.06 0.0095 6 ANKRD9 Uncharacterized protein [Source: UniProtKB / TrEMBL; Acc: F1SA09] 271,000 51.300 -2.4 0.0002 7 APOB Uncharacterized protein [Source: UniProtKB / TrEMBL; Acc: F1SCV9] 122,000 0.000 -6.93 0.0000 8 ATF3 activating transcription factor 3 [Source: HGNC Symbol; Acc: 785] 2,870,000 6,170,000 1.1 0.0051 9 BCAR1 Uncharacterized protein [Source: UniProtKB / TrEMBL; Acc: I3LNL7] 1,700,000 668,000 -1.34 0.0014 10 BCAS1 Sus scrofa breast carcinoma amplified sequence 1 (BCAS1), mRNA. [Source: RefSeq mRNA; Acc: NM_001110175] 761,000 289,000 -1.39 0.0027 11 CD86 Sus scrofa CD86 molecule (CD86), mRNA. [Source: RefSeq mRNA; Acc: NM_214222] 347,000 967,000 1.48 0.0011 12 CDH17 Uncharacterized protein [Source: UniProtKB / TrEMBL; Acc: F1RY44] 228,000 6.030 -5.24 0.0000 13 CDHR2 Uncharacterized protein [Source: UniProtKB / TrEMBL; Acc: F1S3A1] 202,000 6.030 -5.07 0.0000 14 CDHR5 Uncharacterized protein [Source: UniProtKB / TrEMBL; Acc: F1RYX3] 163,000 15.100 -3.44 0.0000 15 CHPF Uncharacterized protein [Source: UniProtKB / TrEMBL; Acc: I3LGL7] 1,810,000 726,000 -1.32 0.0017 16 CHRDL1 Uncharacterized protein [Source: UniProtKB / TrEMBL; Acc: F1RXG9] 53.700 3.020 -4.16 0.0051 17 CHST3 Uncharacterized protein [Source: UniProtKB / TrEMBL; Acc: I3LKW4] 6,170,000 1,240,000 -2.31 0.0000 18 CNN1 Sus scrofa calponin 1, basic, smooth muscle (CNN1), mRNA. [Source: RefSeq mRNA; Acc: NM_213878] 1,990,000 715,000 -1.48 0.0004 19 COL1A2 Sus scrofa collagen, type I, alpha 2 (COL1A2), mRNA. [Source: RefSeq mRNA; Acc: NM_001243655] 57,500,000 28,800,000 -0.998 0.0092 20 COL3A1 Sus scrofa collagen, type III, alpha 1 (COL3A1), mRNA. [Source: RefSeq mRNA; Acc: NM_001243297] 64,800,000 31,600,000 -1.04 0.0068 21 COL4A2 Uncharacterized protein [Source: UniProtKB / TrEMBL; Acc: F1RLL9] 28,600,000 13,900,000 -1.04 0.0068 22 COL5A1 Sus scrofa collagen, type V, alpha 1 (COL5A1), mRNA. [Source: RefSeq mRNA; Acc: NM_001014971] 4,950,000 2,280,000 -1.12 0.0047 23 COL7A1 Uncharacterized protein [Source: UniProtKB / TrEMBL; Acc: F1SKM1] 656,000 257,000 -1.35 0.0045 24 CREB3L3 Uncharacterized protein [Source: UniProtKB / TrEMBL; Acc: F1S7N6] 81.600 9.040 -3.17 0.0037 25 CRNN Sus scrofa cornulin (CRNN), mRNA. [Source: RefSeq mRNA; Acc: NM_001159659] 216,000 25.100 -3.1 0.0000 26 CYP2W1 Uncharacterized protein [Source: UniProtKB / TrEMBL; Acc: F1RIX2] 573,000 95.500 -2.59 0.0000 27 DAGLA Uncharacterized protein [Source: UniProtKB / TrEMBL; Acc: F1RKR2] 523,000 205,000 -1.35 0.0064 28 DAO Sus scrofa D-amino acid oxidase (DAO1), mRNA. [Source: RefSeq mRNA; Acc: NM_214066] 49.800 0.000 -5.63 0.0005 29 DSG1 Sus scrofa desmoglein 1 (DSG1), mRNA. [Source: RefSeq mRNA; Acc: NM_001035535] 165,000 8.040 -4.36 0.0000 30 EMILIN1 Uncharacterized protein [Source: UniProtKB / TrEMBL; Acc: F1SDQ5] 1,280,000 410,000 -1.65 0.0002 31 EPS8L3 Uncharacterized protein [Source: UniProtKB / TrEMBL; Acc: F1S606] 73.600 0.000 -6.19 0.0000 32 EVPL Uncharacterized protein [Source: UniProtKB / TrEMBL; Acc: F1RW07] 7,740,000 3,380,000 -1.2 0.0023 33 F10 Sus scrofa coagulation factor X protein (LOC733662), mRNA. [Source: RefSeq mRNA; Acc: NM_001044592] 71.600 2.010 -5.16 0.0003 34 FAM174B Uncharacterized protein [Source: UniProtKB / TrEMBL; Acc: F1SA78] 179,000 37.200 -2.27 0.0017 35 FAM89A Uncharacterized protein [Source: UniProtKB / TrEMBL; Acc: F1RGS7] 488,000 195,000 -1.32 0.0085 36 FBLN5 Uncharacterized protein [Source: UniProtKB / TrEMBL; Acc: F1SD87] 271,000 59.300 -2.19 0.0005 37 FBP2 Sus scrofa fructose-1,6-bisphosphatase 2 (FBP2), mRNA. [Source: RefSeq mRNA; Acc: NM_001167632] 160,000 24.100 -2.73 0.0005 38 FHL1C Sus scrofa four and a half LIM domains 1 protein, isoform C (FHL1C), mRNA. [Source: RefSeq mRNA; Acc: NM_214375] 1,580,000 612,000 -1.37 0.0013 39 GJB1 gap junction protein, beta 1, 32 kDa [Source: HGNC Symbol; Acc: 4283] 942,000 393,000 -1.26 0.0048 40 GLDN Uncharacterized protein [Source: UniProtKB / TrEMBL; Acc: F1RYN3] 280,000 66.300 -2.08 0.0007 41 GPR153 Uncharacterized protein [Source: UniProtKB / TrEMBL; Acc: F1RIL9] 1,270,000 585,000 -1.12 0.0092 42 HAL Histidine ammonia-lyase [Source: UniProtKB / TrEMBL; Acc: F1SQR7] 428,000 78.400 -2.45 0.0000 43 HNF4A Sus scrofa hepatocyte nuclear factor 4, alpha (HNF4A), mRNA. [Source: RefSeq mRNA; Acc: NM_001044571] 131,000 0.000 -7.03 0.0000 44 IGFBP5 Uncharacterized protein [Source: UniProtKB / TrEMBL; Acc: I3LJI8] 840,000 314,000 -1.42 0.0020 45 IGSF23 Uncharacterized protein [Source: UniProtKB / TrEMBL; Acc: F1RMS9] 155,000 0.000 -7.27 0.0000 46 IL21R interleukin 21 receptor [Source: HGNC Symbol; Acc: 6006] 348,000 92.500 -1.91 0.0008 47 ITIH4 inter-alpha-trypsin inhibitor heavy chain H4 [Source: RefSeq peptide; Acc: NP_001001537] 573,000 18.100 -4.99 0.0000 48 KCNK5 Uncharacterized protein [Source: UniProtKB / TrEMBL; Acc: F1RVQ9] 1,900,000 781,000 -1.28 0.0021 49 KRT1 Uncharacterized protein [Source: UniProtKB / TrEMBL; Acc: F1SGG3] 2,440,000 2.010 -10.2 0.0000 50 KRT13 Uncharacterized protein [Source: UniProtKB / TrEMBL; Acc: F1S0K1] 2,630,000 548,000 -2.26 0.0000 51 KRT14 Uncharacterized protein [Source: UniProtKB / TrEMBL; Acc: F1S0L1] 944,000 66.300 -3.83 0.0000 52 KRT15 Uncharacterized protein [Source: UniProtKB / TrEMBL; Acc: F1S0K2] 148,000 2.010 -6.2 0.0000 53 KRT20 Keratin, type I cytoskeletal 20 [Source: UniProtKB / Swiss-Prot; Acc: Q29218] 37.800 1,000 -5.23 0.0097 54 KRT4 Uncharacterized protein [Source: UniProtKB / TrEMBL; Acc: I3LQN8] 2,930,000 89.400 -5.03 0.0000 55 KRT5 Uncharacterized protein [Source: UniProtKB / TrEMBL; Acc: F1SGG6] 3,240,000 44.200 -6.19 0.0000 56 KRT77 Uncharacterized protein [Source: UniProtKB / TrEMBL; Acc: I3LNT6] 84.600 0.000 -6.39 0.0000 57 LCN15 Uncharacterized protein [Source: UniProtKB / TrEMBL; Acc: F1RVY2] 32.800 0.000 -5.03 0.0070 58 LGALS2 Sus scrofa lectin, galactoside-binding, soluble, 3 (LGALS3), mRNA. [Source: RefSeq mRNA; Acc: NM_001142842] 145,000 0.000 -7.18 0.0000 59 LOX lysyl oxidase [Source: RefSeq peptide; Acc: NP_001193332] 628,000 168,000 -1.9 0.0001 60 LREAP1 Sus scrofa low density lipoprotein receptor-related protein associated protein 1 (LRPAP1), mRNA. [Source: RefSeq mRNA; Acc: NM_001113436] 5,730,000 2,820,000 -1.02 0.0093 61 LTBP2 Uncharacterized protein [Source: UniProtKB / TrEMBL; Acc: F1S2T5] 1,470,000 383,000 -1.94 0.0000 62 MAMDC4 Uncharacterized protein [Source: UniProtKB / TrEMBL; Acc: F1RW01] 925,000 77.400 -3.58 0.0000 63 MAOB Sus scrofa monoamine oxidase B (MAOB), nuclear gene encoding mitochondrial protein, mRNA. [Source: RefSeq mRNA; Acc: NM_001001864] 254,000 58.300 -2.12 0.0008 64 MFSD10 Uncharacterized protein [Source: UniProtKB / TrEMBL; Acc: F1S8P6] 588,000 246,000 -1.26 0.0092 65 MMP8 Uncharacterized protein [Source: UniProtKB / TrEMBL; Acc: F1SV69] 91.500 11.100 -3.05 0.0029 66 MPV17L2 Uncharacterized protein [Source: UniProtKB / TrEMBL; Acc: F1S919] 320,000 114,000 -1.5 0.0078 67 MUC13A Sus scrofa mucin 13, cell surface associated (MUC13), mRNA. [Source: RefSeq mRNA; Acc: NM_001105293] 55.700 0.000 -5.79 0.0002 68 MUC4 Sus scrofa mucin 4, cell surface associated (MUC4), mRNA. [Source: RefSeq mRNA; Acc: NM_001206344] 3,790,000 1,600,000 -1.25 0.0019 69 MXRA5 Uncharacterized protein [Source: UniProtKB / TrEMBL; Acc: F1RZ06] 19,900,000 9,410,000 -1.08 0.0050 70 NPC1L1 Uncharacterized protein [Source: UniProtKB / TrEMBL; Acc: F1SSH5] 68.700 1,000 -6.09 0.0002 71 PAPSS2 3'-phosphoadenosine 5'-phosphosulfate synthase 2 [Source: HGNC Symbol; Acc: 8604] 7,260,000 1,680,000 -2.11 0.0000 72 PCDH1 Uncharacterized protein [Source: UniProtKB / TrEMBL; Acc: I3L7J4] 3,080,000 1,450,000 -1.09 0.0070 73 PCK1 Sus scrofa phosphoenolpyruvate carboxykinase 1 (soluble) (PCK1), mRNA. [Source: RefSeq mRNA; Acc: NM_001123158] 156,000 14.100 -3.47 0.0000 74 PIM3 pim-3 oncogene [Source: HGNC Symbol; Acc: 19310] 466,000 41.200 -3.5 0.0000 75 PPL Uncharacterized protein [Source: UniProtKB / TrEMBL; Acc: F1RK90] 5,930,000 2,500,000 -1.25 0.0016 76 PPP1R3D Protein phosphatase 1 regulatory subunit 3 [Source: UniprotKB / TrEMBL; Acc: F1RJ12] 510,000 140,000 -1.87 0.0003 77 PRAP1 Uncharacterized protein [Source: UniProtKB / TrEMBL; Acc: F1SAB9] 46.800 1,000 -5.54 0.0029 78 PRKCDBP Uncharacterized protein [Source: UniProtKB / TrEMBL; Acc: F1RMM0] 577,000 65.300 -3.14 0.0000 79 PRKG2 cGMP-dependent protein kinase [Source: UniProtKB / TrEMBL; Acc: F1RVC8] 235,000 37.200 -2.66 0.0001 80 RBP2 Sus scrofa retinol binding protein 2, cellular (RBP2), mRNA. [Source: RefSeq mRNA; Acc: NM_214451] 192,000 0.000 -7.58 0.0000 81 RBP4 Sus scrofa retinol binding protein 4, plasma (RBP4), mRNA. [Source: RefSeq mRNA; Acc: NM_214057] 729,000 306,000 -1.26 0.0069 82 REG4 Sus scrofa regenerating islet-derived family, member 4 (REG4), mRNA. [Source: RefSeq mRNA; Acc: NM_001190251] 66.700 0.000 -6.05 0.0000 83 RIPK4 Uncharacterized protein [Source: UniProtKB / TrEMBL; Acc: F1SG49] 3,120,000 1,500,000 -1.05 0.0087 84 ROS1 Uncharacterized protein [Source: UniProtKB / TrEMBL; Acc: I3LEW1] 46.800 1,000 -5.54 0.0029 85 SCARA5 Uncharacterized protein [Source: UniProtKB / TrEMBL; Acc: F1RJR8] 456,000 151,000 -1.6 0.0021 86 SDK1 Uncharacterized protein [Source: UniProtKB / TrEMBL; Acc: F1RI55] 310,000 83.400 -1.9 0.0012 87 SH3GL2 Uncharacterized protein [Source: UniProtKB / TrEMBL; Acc: F1SNE5] 274,000 90.500 -1.6 0.0071 88 SLC15A1 Sus scrofa solute carrier family 15 (oligopeptide transporter), member 1 (SLC15A1), mRNA. [Source: RefSeq mRNA; Acc: NM_214347] 4,680,000 2,060,000 -1.18 0.0029 89 SLC16A3 Uncharacterized protein [Source: UniProtKB / TrEMBL; Acc: I3LQ25] 214,000 51.300 -2.06 0.0020 90 SLC2A8 Uncharacterized protein [Source: UniProtKB / TrEMBL; Acc: F1RS19] 709,000 291,000 -1.28 0.0061 91 SLC38A10 Uncharacterized protein [Source: UniProtKB / TrEMBL; Acc: I3LB54] 1,650,000 713,000 -1.21 0.0041 92 SLC45A3 Uncharacterized protein [Source: UniProtKB / TrEMBL; Acc: I3L8D4] 4,850,000 865,000 -2.49 0.0000 93 SLC45A4 Uncharacterized protein [Source: UniProtKB / TrEMBL; Acc: F1RSK5] 1,570,000 530,000 -1.57 0.0003 94 SLC5A1 Sus scrofa solute carrier family 5 (sodium / glucose cotransporter), member 1 (SLC5A1), mRNA. [Source: RefSeq mRNA; Acc: NM_001164021] 193,000 14.100 -3.78 0.0000 95 SLC5A6 Uncharacterized protein [Source: UniProtKB / TrEMBL; Acc: F1SED7] 437,000 164,000 -1.41 0.0063 96 SLC6A19 Transporter [Source: UniProtKB / TrEMBL; Acc: F1S025] 100,000 0.000 -6.64 0.0000 97 SLIT3 Uncharacterized protein [Source: UniProtKB / TrEMBL; Acc: F1RR92] 1,360,000 537,000 -1.34 0.0018 98 SPOCK3 Uncharacterized protein [Source: UniProtKB / TrEMBL; Acc: F1RIW2] 92.500 10.100 -3.2 0.0020 99 SYNM Uncharacterized protein [Source: UniProtKB / TrEMBL; Acc: I3LKP0] 692,000 165,000 -2.07 0.0000 100 THBS2 Uncharacterized protein [Source: UniProtKB / TrEMBL; Acc: F1SBZ9] 1,440,000 630,000 -1.2 0.0050 101 TM4SF20 Uncharacterized protein [Source: UniProtKB / TrEMBL; Acc: F1SNM8] 200,000 0.000 -7.64 0.0000 102 TOR4A Uncharacterized protein [Source: UniProtKB / TrEMBL; Acc: F1RVY0] 79.600 7.040 -3.5 0.0022 103 TRIM3 Uncharacterized protein [Source: UniProtKB / TrEMBL; Acc: F1RMN8] 434,000 165,000 -1.4 0.0070 104 UPK1A Sus scrofa uroplakin 1A (UPK1A), mRNA. [Source: RefSeq mRNA; Acc: NM_001123211] 187,000 20.100 -3.22 0.0000 105 USH1C Uncharacterized protein [Source: UniProtKB / TrEMBL; Acc: F1S9A7] 31.800 0.000 -4.99 0.0081 106 VAT1L Uncharacterized protein [Source: UniProtKB / TrEMBL; Acc: F1S467] 102,000 20.100 -2.35 0.0097 107 VIL1 Villin-1 [Source: UniProtKB / TrEMBL; Acc: F1SRY3] 219,000 12.100 -4.18 0.0000 108 ZFYVE21 Uncharacterized protein [Source: UniProtKB / TrEMBL; Acc: F1S9X1] 101,000 19.100 -2.41 0.0086

While the present invention has been particularly shown and described with reference to specific embodiments thereof, those skilled in the art will appreciate that such specific embodiments are merely preferred embodiments and that the scope of the present invention is not limited thereby. something to do. It is therefore intended that the scope of the invention be defined by the claims appended hereto and their equivalents.

Claims (5)

Pig gene CHI3L1, EGR2, HBA, LIPG, NFKBIZ, PDPN, PHEROC, PLAT, PLAUR, PPP1R15A, SPRY2, XIRP1, ZNF385D, A2ML1, ACTG2, ADAMTS8, ALDH1A2, ANKH, ANKRD9, APOB, ATF3, BCAR1, BCAS1, CDL17, CDHR2, CDHR5, CHPF, CHRDL1, CHST3, CNN1, COL1A2, COL3A1, COL4A2, COL5A1, COL7A1, CREB3L3, CRNN, CYP2W1, DAGLA, DAO, DSG1, EMILIN1, EPS8L3, EVPL, F10, FAM174B, FAM89A, FBLN5, IGFBP5, IGSF23, IL21R, ITIH4, KCNK5, KRT1, KRT13, KRT14, KRT15, KRT20, KRT4, KRT5, KRT77, LCN15, LGALS2, LOX, LREAP1, LTBP2, LBP2, FHL1C, GJB1, GLDN, GPR153, HAL, HNF4A, MAKD, MAMDC4, MAOB, MFSD10, MMP8, MPV17L2, MUC13A, MUC4, MXRA5, NPC1L1, PAPSS2, PCDH1, PCK1, PIM3, PPL, PPP1R3D, PRAP1, PRKCDBP, PRKG2, RBP2, RBP4, REG4, RIPK4, ROS1, SCARA5, SDK1, 1 selected from the group consisting of SH3GL2, SLC15A1, SLC16A3, SLC2A8, SLC38A10, SLC45A3, SLC45A4, SLC5A1, SLC5A6, SLC6A19, SLIT3, SPOCK3, SYNM, THBS2, TM4SF20, TOR4A, TRIM3, UPK1A, USH1C, VAT1L, VIL1 and ZFYVE21 There is a need for an agent that measures the expression level of abnormal genes. Wherein the composition is used for predicting the number of pigs. The composition according to claim 1, wherein the number of hatchlings of black pig is predicted. A kit for estimating the number of pigs of a pig comprising the composition according to claim 1 or 2. 4. The kit according to claim 3, wherein the kit is an RT-PCR kit, a microarray chip kit or a protein chip kit. Extracting mRNA from two or more pigs, quantifying the expression level of each gene, and obtaining an average expression level; And
Wherein at least one gene of CHI3L1, EGR2, HBA, LIPG, NFKBIZ, PDPN, PHEROC, PLAT, PLAUR, PPP1R15A, SPRY2, XIRP1 and ZNF385D is expressed higher than the average expression amount in each quantified gene, CYP2W1, CYP2W1, CYP2W1, CYP2W1, CYP2W1, CYP2W1, CYP2W1, CYP2W1, CYP2W1, DAGLA, DAO, DSG1, EMILIN1, EPS8L3, EVPL, F10, FAM174B, FAM89A, FBLN5, FBP2, FHL1C, GJB1, GLDN, GPR153, HAL, HNF4A, IGFBP5, IGSF23, IL21R, ITIH4, KCNK5, KRT1, KRT13, KRT14, MCPD1, MMP8, MPV17L2, MUC13A, MUC4, MXRA5, NPC1L1, PAPSS2, PCDH1, PCK1, PIM3, PPL, PPP1R3D, MKD15, KRT20, KRT4, KRT5, KRT77, LCN15, LGALS2, LOX, LREAP1, LTBP2, MAMDC4, MAOB, PRP1, PRKCDBP, PRKG2, RBP2, RBP4, REG4, RIPK4, ROS1, SCARA5, SDK1, SH3GL2, SLC15A1, SLC16A3, SLC2A8, SLC38A10, SLC45A3, SLC45A4, SLC5A1, SLC5A6, SLC6A19, SLIT3, SPOCK3, SYNM, THBS2, TM4SF20, TOR4A, TRIM3, UPK1A, USH1C, VAT1L, VIL 1 < / RTI > and ZFYVE21 is expressed in an amount less than the average expression level, is predicted to a pig having a higher number of pigs.

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