A SNP molecular marker associated with total litter size in pigs and its application

Abstract

This invention provides a SNP molecular marker associated with the total litter size trait in pigs and its application, relating to the field of biotechnology. The application includes using substances detecting SNP sites in the preparation of products predicting the total litter size trait in pigs. The SNP sites include one or more of SNP sites rs334299237, rs318687930, and rs325934988. These SNP sites are associated with the total litter size trait in pigs and can be used to predict this trait and for the screening and breeding of sows with high total litter size.

Description

Technical Field

[0001] This invention relates to the field of biotechnology, and in particular to a SNP molecular marker related to the trait of total litter size in pigs and its application. Background Technology

[0002] Reproductive traits in pigs are important indicators for measuring the reproductive capacity of sows. These mainly include litter size, piglet birth weight, number of teats, milk production (litter weight at 21 days of age), and the number of piglets recovered at weaning. These traits are extremely important for pig farming. Good reproductive performance in sows means that each sow can produce more healthy piglets within a certain period, reducing breeding costs and improving production efficiency.

[0003] Total litter size is of great significance for assessing sow reproductive efficiency and pig breeding. By studying total litter size, breeding pigs with high litter size potential can be screened, the transmission and fixation of superior genes can be accelerated, the reproductive capacity of pig herds can be improved, and a scientific basis can be provided for breeding pig selection and genetic improvement.

[0004] Therefore, the identification of genes related to total litter size in pigs can provide important clues for explaining the genetic mechanisms of fetal growth and development in pigs and other mammals, and provide a theoretical basis for the genetic improvement of reproductive traits in pigs.

[0005] In view of this, the present invention is hereby proposed. Summary of the Invention

[0006] The purpose of this invention is to overcome the shortcomings of the prior art and provide an SNP molecular marker related to the total number of piglets born and its application.

[0007] To solve the above-mentioned technical problems, the present invention adopts the following technical solution:

[0008] In a first aspect, a substance for detecting SNP sites is provided for use in the preparation of products that predict the total number of piglets produced, wherein the SNP sites include one or more of the following: SNP site rs334299237, SNP site rs318687930 and SNP site rs325934988.

[0009] Secondly, a kit for predicting the total litter size trait in pigs is provided, the kit comprising at least one of the following primer pairs:

[0010] The primer pairs used to amplify SNP site rs334299237 have nucleotide sequences as shown in SEQ ID NO. 1 and 2, respectively; the primer pairs used to amplify SNP site rs318687930 have nucleotide sequences as shown in SEQ ID NO. 3 and 4, respectively; and the primer pairs used to amplify SNP site rs325934988 have nucleotide sequences as shown in SEQ ID NO. 5 and 6, respectively.

[0011] Thirdly, a method for predicting the total litter size trait in pigs is provided, the method comprising predicting sows that meet at least one of (i) to (iii) as sows with a high total litter size trait:

[0012] (i) The genotype of SNP locus rs334299237 is TC.

[0013] (ii) The genotype of SNP locus rs318687930 is GA;

[0014] (iii) The genotype of SNP locus rs325934988 is GA.

[0015] Fourthly, a method for screening sows with a high total litter size trait is provided, the method comprising screening sows with a high total litter size trait predicted by the method described in the third aspect.

[0016] Fifthly, a breeding method for sows exhibiting a high total litter size trait is provided, the method comprising acquiring at least one of the following genotypes in the sow:

[0017] (i) The genotype of SNP locus rs334299237 is TC.

[0018] (ii) The genotype of SNP locus rs318687930 is GA;

[0019] (iii) The genotype of SNP locus rs325934988 is GA.

[0020] Compared with the prior art, the present invention has the following beneficial effects:

[0021] This invention utilizes low-depth resequencing and genome-wide association analysis to identify SNP molecular markers associated with the total litter size trait in pigs. Three SNP loci, rs334299237, rs318687930, and rs325934988, were found to be associated with this trait. Sows with specific genotypes at these three loci exhibit higher total litter size. These SNP molecular markers can be used to predict total litter size, screen sows with high total litter size, and develop breeding methods to obtain sows with high total litter size. The application of these SNP loci helps improve sow reproductive performance, bringing greater economic benefits to the pig farming industry. Attached Figure Description

[0022] To more clearly illustrate the specific embodiments of the present invention or the technical solutions in the prior art, the drawings used in the description of the specific embodiments or the prior art will be briefly introduced below. Obviously, the drawings described below are some embodiments of the present invention. For those skilled in the art, other drawings can be obtained from these drawings without creative effort.

[0023] Figure 1 This is a genotype detection peak diagram obtained by capillary electrophoresis in an embodiment of the present invention. Detailed Implementation

[0024] The following will clearly and completely describe the technical solutions of the present invention in conjunction with the embodiments. Obviously, the embodiments described are only some embodiments of the present invention, not all embodiments. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without making creative efforts are within the scope of protection of the present invention.

[0025] cAMP response element binding protein 3 (CREB3 regulatory factor, CREBRF), also known as the Luman / CREB3 recruiting factor (LRF), is a regulator of the Luman / CREB3 protein (cyclic adenosinemonophosphate response element binding protein 3, Luman / CREB3). Studies have shown that CREBRF knockout mice have significantly lower prolactin levels than normal mice, impaired glucocorticoid signaling, and exhibit severe maternal behavioral defects, lacking the instinct to care for offspring. CREBRF gene expression is high in the ovary and uterus during estrus and postestrus, suggesting that CREBRF may be involved in cell differentiation, embryonic development, and implantation. Another study showed that CREBRF can mediate ERS pathway-mediated apoptosis of GCs and plays a crucial role in mouse follicle selection. CREBRF expression can be inhibited by E2 but induced by P4, and CREBRF can regulate decidualization during pregnancy by modulating the proliferation of mouse uterine stromal cells. Therefore, the CREBRF gene plays an important regulatory role in animal growth, development, and reproduction, especially in follicle growth and development, cell differentiation, embryonic development, and implantation. Furthermore, polymorphic changes at CREBRF gene SNP sites are correlated with estrogen levels, which in turn regulate sow reproductive efficiency. Total litter size is an important indicator of sow reproductive traits; therefore, this invention investigates the correlation between CREBRF gene single nucleotide polymorphisms and total litter size.

[0026] This invention employs low-depth resequencing and genome-wide association analysis to analyze the association between CREBRF gene polymorphisms and important reproductive traits, and identifies SNP molecular markers significantly associated with the total litter size trait in the pig genome. This invention discovered three SNP loci, rs334299237, rs318687930, and rs325934988, that are associated with the total litter size trait in pigs.

[0027] The CREBRF gene SNP site rs334299237 is located at nucleotide position 51160002 on pig chromosome 16 (reference genome is Sscrofa11.1). When the genotype of this SNP site is TC, that is, when the nucleotide position 51160002 on pig chromosome 16 is a heterozygote of T and C, the sow has a high total litter size at first birth.

[0028] The CREBRF gene SNP site rs318687930 is located at nucleotide position 51174074 on pig chromosome 16 (reference genome is Sscrofa11.1). When the genotype of this SNP site is GA, that is, nucleotide position 51174074 on pig chromosome 16 is a heterozygote of G and A, the sow has a high total litter size trait.

[0029] The CREBRF gene SNP site rs325934988 is located at nucleotide position 51225561 on chromosome 16 of pigs (reference genome is Sscrofa11.1). When the genotype of this SNP site is GA, that is, the nucleotide position 51225561 of the pig is a heterozygote of G and A, the sow has a high total litter size at first birth.

[0030] In this article, "total primiparous litter size" refers to the total number of piglets born to a sow in her first litter, including live piglets and stillborn piglets. "High primiparous litter size" refers to a total primiparous litter size that is higher than the average total primiparous litter size for that breed. In an optional implementation, when the pig breed is Large White, "high primiparous litter size" is defined as a total primiparous litter size of at least 12 piglets.

[0031] In this article, "total multiparous litter size" refers to the average number of piglets born in each litter from the second to the nth litter, where n is a positive integer ≥2. The total number of piglets born in each litter includes live and stillborn piglets. "High multiparous litter size" refers to a total multiparous litter size that is higher than the average multiparous litter size for that breed. In an optional implementation, when the pig breed is Large White, "high multiparous litter size" is a total multiparous litter size of at least 14 piglets.

[0032] In this article, "total litter size" includes at least one of the traits defined above as "primiparous total litter size" and "multiparous total litter size"; "high total litter size" includes at least one of the traits defined above as "high primiparous total litter size" and "high multiparous total litter size".

[0033] Based on the above findings, in a first aspect, an application is provided for a substance for detecting SNP sites in the preparation of a product for predicting the total number of piglets produced, wherein the SNP sites include one or more of the following: SNP site rs334299237, SNP site rs318687930 and SNP site rs325934988.

[0034] In an optional implementation, the total number of piglets born includes the total number of primiparous piglets and / or the total number of multiparous piglets.

[0035] In an optional embodiment, the product includes a substance for detecting the SNP site rs334299237, and the total litter size trait of pigs includes the total litter size of primiparous pigs.

[0036] In an optional embodiment, the product includes a substance for detecting the SNP locus rs318687930, and the total litter size trait of pigs includes primiparous total litter size and multiparous total litter size.

[0037] In an optional embodiment, the product includes a substance for detecting the SNP locus rs325934988, and the total litter size trait of pigs includes the total litter size of primiparous pigs.

[0038] In an optional embodiment, the product includes a substance that detects SNP sites rs334299237, rs318687930, and rs325934988.

[0039] In optional embodiments, the substances used to detect SNP sites include, but are not limited to, one or more of the following: reagents for nucleic acid amplification, reagents for detecting nucleic acid amplification products, reagents for constructing sequencing libraries, and reagents for sequencing. More specifically, the substances used to detect SNP sites include, but are not limited to, primers, probes, enzymes for nucleic acid amplification reactions, fluorescent labels, buffer reagents, dNTPs, salts, etc. Depending on the specific detection method, those skilled in the art can select the above-mentioned reagents for nucleic acid amplification, reagents for detecting nucleic acid amplification products, reagents for constructing sequencing libraries, and reagents for sequencing according to the methods described in general and more specific textbooks, references, process manuals, product instructions, and standard documents. This invention does not impose any limitations on these selections.

[0040] In an optional embodiment, the substance for detecting SNP sites includes at least one of the following primer pairs: a primer pair for amplifying SNP site rs334299237, with nucleotide sequences as shown in SEQ ID NO. 1 and 2, respectively; a primer pair for amplifying SNP site rs318687930, with nucleotide sequences as shown in SEQ ID NO. 3 and 4, respectively; and a primer pair for amplifying SNP site rs325934988, with nucleotide sequences as shown in SEQ ID NO. 5 and 6, respectively.

[0041] Secondly, a kit for predicting the total litter size trait in pigs is provided, the kit comprising at least one of the following primer pairs: a primer pair for amplifying SNP site rs334299237, with nucleotide sequences as shown in SEQ ID NO. 1 and 2, respectively; a primer pair for amplifying SNP site rs318687930, with nucleotide sequences as shown in SEQ ID NO. 3 and 4, respectively; and a primer pair for amplifying SNP site rs325934988, with nucleotide sequences as shown in SEQ ID NO. 5 and 6, respectively.

[0042] Thirdly, a method for predicting the total litter size trait in pigs is provided, the method comprising predicting sows that meet at least one of (i) to (iii) as sows with a high total litter size trait:

[0043] (i) The genotype of SNP locus rs334299237 is TC.

[0044] (ii) The genotype of SNP locus rs318687930 is GA;

[0045] (iii) The genotype of SNP locus rs325934988 is GA.

[0046] In an optional implementation, the method includes predicting sows that meet the criteria of (i) to (iii) above as sows with a high total litter size trait.

[0047] In an optional implementation, the method for predicting the total number of piglets produced includes predicting sows that meet at least one of (i) and / or (iii) as sows with a high initial total number of piglets produced trait:

[0048] (i) The genotype of SNP locus rs334299237 is TC.

[0049] (iii) The genotype of SNP locus rs325934988 is GA.

[0050] In an optional implementation, the method for predicting the total number of piglets produced includes predicting sows that meet (ii) as sows with high primiparous total number of piglets produced and high multiparous total number of piglets produced:

[0051] (ii) The genotype of SNP site rs318687930 is GA.

[0052] The third aspect provides a method for predicting the total litter size trait in pigs, which includes using one or more of the SNP loci rs334299237, rs318687930, and rs325934988 as identification targets; it can also be combined with other targets, such as other polymorphic molecular markers (e.g., SSR molecular markers, STR molecular markers, or InDel molecular markers), or other screening methods known in the art to assist in the prediction of the total litter size trait in pigs.

[0053] Fourthly, a method for screening sows with a high total litter size trait is provided, comprising screening sows with a high total litter size trait predicted by the method described in the third aspect.

[0054] Fifthly, a breeding method for sows exhibiting a high total litter size trait is provided, the breeding method comprising obtaining at least one of the following genotypes in the sow:

[0055] (i) The genotype of SNP locus rs334299237 is TC.

[0056] (ii) The genotype of SNP locus rs318687930 is GA;

[0057] (iii) The genotype of SNP locus rs325934988 is GA.

[0058] In an optional embodiment, the breeding method includes enabling sows to acquire the genotype of SNP locus rs334299237 as TC, in order to increase the total number of piglets born in the first litter of sows.

[0059] In an optional embodiment, the breeding method includes enabling sows to acquire the genotype of GA at the SNP locus rs318687930, in order to increase the total number of piglets in primiparous litters and the total number of piglets in multiparous litters.

[0060] In an optional embodiment, the breeding method includes enabling sows to acquire the genotype of GA at the SNP locus rs325934988, in order to increase the total number of piglets born in the first litter.

[0061] The specific means by which a sow acquires a specific genotype can be implemented by those skilled in the art using conventional methods known in the art, and this invention does not limit this. In an optional embodiment, offspring with the target genotype are obtained by acquiring parents with a known genotype. In another optional embodiment, offspring with the target genotype are obtained through gene editing methods.

[0062] In an optional implementation, the pig in any of the above implementations is a Large White pig.

[0063] The present invention will be further illustrated below with specific embodiments. However, it should be understood that these embodiments are merely for the purpose of more detailed illustration and should not be construed as limiting the present invention in any way.

[0064] Example 1

[0065] I. Sample Collection

[0066] The experimental population in this embodiment consisted of 226 British Large White pigs from Rizhao Breeding Farm in Shandong, 165 Canadian Large White pigs from Xinjiang Tiankang Jiamei Breeding Branch, and 199 Danish Large White pigs from the great-grandparent breeding farm of Xinjiang Tiankang Animal Husbandry Technology Co., Ltd. A total of 590 Large White pigs were used for CREBRF gene SNP genotyping. Ear tissue samples were collected from each pig, placed in 75% alcohol, and stored at -20°C for DNA extraction.

[0067] II. Extraction and Library Construction of Pig Genomic DNA

[0068] (1) Cut an appropriate amount of pig ear tissue, cut it into pieces and put it into a 1.5mL Axgen tube.

[0069] (2) Take a 50mL BD centrifuge tube, mix proteinase K and lysis buffer to a final concentration of 0.4mg / mL evenly, and add 0.5mL lysis buffer to a 1.5mL centrifuge tube containing pig ear tissue for lysis.

[0070] (3) Place the centrifuge tubes evenly and parallel on the shaking plate of the constant temperature hybridization oven (tightly seal the tube caps to prevent liquid leakage). Incubate at 55°C for more than 6 hours (thorough mixing of the sample during digestion is very important. The criteria for judgment are the absence of obvious visible pig ear tissue and the milky consistency of the digested mixture).

[0071] (4) After the sample has been fully lysed, remove the centrifuge tubes, add 0.3 mL of saturated sodium chloride solution to each tube, mix thoroughly by inverting 6-8 times, and then place them on ice for 15 minutes.

[0072] (5) After the ice bath, centrifuge at 12,000 rpm at room temperature for 15 minutes. Carefully and slowly transfer the supernatant to a new 1.5 mL Axgen centrifuge tube (be careful to avoid pouring out the precipitate along with the supernatant, keep the pouring method consistent, and keep the amount of supernatant poured out in each tube the same).

[0073] (6) Add 0.7 mL of isopropanol (the amount of isopropanol added varies with the amount of supernatant poured out, and the two are equal in volume) to each tube, and invert until flocculent precipitate appears in the solution (if no flocculent precipitate appears, the solution can be placed in a -20°C refrigerator for 2 hours or in a 4°C refrigerator overnight).

[0074] (7) Centrifuge at 12000 rpm at room temperature for 15 minutes and remove the supernatant.

[0075] (8) Add 0.5 mL of 70% ethanol to each centrifuge tube and gently invert to thoroughly rinse the DNA that has precipitated above.

[0076] (9) Centrifuge at 10000 rpm for 30 seconds, then use a 200 μL micropipette to remove the ethanol from the centrifuge tube, leaving the precipitated DNA in the tube.

[0077] (10) Air dry the DNA for 10 minutes.

[0078] (11) Pipette 0.1 mL of TE buffer into each tube to redissolve the DNA precipitate. Place the tube at 55°C for 2 hours and shake it several times to ensure that the DNA is fully dissolved.

[0079] (12) After the DNA was fully dissolved, the concentration of the extracted DNA was determined by UV spectrophotometer, and the quality of the extracted DNA was detected by agarose gel electrophoresis. The quality of the DNA was further tested using Qbuit, and qualified samples were used for library construction.

[0080] (13) Library construction: Sequencing libraries were prepared using the BGI library construction kit.

[0081] (14) Sequencing: After the library preparation is completed and the quality is checked, PE150 mode sequencing is performed using DNBSEQ-T7.

[0082] III. SNP chip genotyping and genotypic data quality control

[0083] DNA samples from 590 Large White pigs were sent to Beijing Gezhi Boya Biotechnology Co., Ltd. for 20× depth sequencing and low-depth resequencing.

[0084] Low-quality sequencing data was removed using the default parameters of fastp v0.23.2 to obtain clean reads. The clean reads were then aligned to the reference genome using the default parameters of bwa-mem v0.7.17. The alignment results were then converted into BAM files and sorted using Samtools software. Picard MarkDuplicates was used to remove duplicate reads from the sorted BAM files. Genomic variant detection was performed using GATK-HaplotypeCaller software, generating gvcf files for individual samples. GATK CombineGVCFs and GenotypeGVCFs were then used to merge and genotype the gvcf files. The filtering parameters for SNPs and Indels are shown in Table 1.

[0085] Table 1 Filtering Parameters

[0086] Filtering parameters SNP Indel QD <2.0 <2.0 QUAL <30.0 <30.0 MQ <40.0 <40.0 SOR >3.0 FS >60.0 >200.0 MQRankSum <-12.5 ReadPosRankSum <-8.0 <-20.0

[0087] IV. Data Processing and Analysis

[0088] (1) Phenotypic data analysis: The phenotypic data of total litter size of 590 Large White pigs collected were organized in Excel. Descriptive statistics were performed on the phenotypic data of total litter size, and the sample size, mean, standard deviation, standard error, maximum value, minimum value and quartiles of each variable were calculated.

[0089] (2) Statistical Analysis: Excel 2021 was used to calculate various polymorphism indicators. The relevant formulas for calculating polymorphism indicators are as follows:

[0090] (2.1) Gene frequency (AF)

[0091]

[0092] (2.2) Genotype frequency (GF)

[0093]

[0094] (2.3) Genetic heterozygosity (He)

[0095]

[0096] (2.4) Effective number of alleles (Ne)

[0097]

[0098] (2.5) Polymorphic Information Content (PIC)

[0099]

[0100] (2.6) Correlation Analysis

[0101] Anova analysis was performed using the LSD method. Reproductive phenotypic characteristics of different genotypes were expressed as mean ± standard deviation. A linear model using PLINK software was used to perform association analysis between genotype and reproductive phenotype. The field effect was used as a covariate for correction. P ≤ 0.05 was considered statistically significant, and P ≤ 0.01 was considered extremely statistically significant.

[0102] V. Capillary electrophoresis detection of SNPs

[0103] (1) Determine the number of reactions N. During the test, a template-free control (nuclease-free pure water) should be tested simultaneously. N = number of samples to be tested (n) + template-free control (1) + 1.

[0104] (2) Place the enzyme mixture on ice, and thaw the remaining components at room temperature until completely melted. Mix with a vortex mixer for 10 seconds, and then centrifuge with a micro centrifuge for 10 seconds to concentrate the liquid at the bottom of the tube.

[0105] (3) Prepare the reaction system except for the template according to Table 2. Take one centrifuge tube, add each component, mix with a vortex mixer for 10s, centrifuge with a microcentrifuge for 10s, and aliquot 9μL into PCR reaction tubes. Primer sequences are shown in Table 3.

[0106] Table 2 Reaction system configuration

[0107] Serial Number Component Name Amount of standard system added (μL) Reaction number N and amount added (μL) 1 2.5×PCR reaction solution 4 4N 2 primer mixture 1 1N 3 Nuclease-free water 4 4N total 9 9N

[0108] Note: PCR products should be stored at 2–8℃ and tested within 24 hours, or stored at -20℃±5℃ and tested within 7 days. The number of freeze-thaw cycles should not exceed 3.

[0109] Table 3 Primer sequences

[0110]

[0111] (4) Add 1 μL of the sample to be tested to each of the pre-dispensed reaction tubes. Record the order of the numbers, mix using a vortex mixer for 10 s, and then centrifuge using a microcentrifuge for 10 s.

[0112] (5) Place each reaction tube in the reaction chamber of the PCR instrument and run the PCR program. The PCR amplification program is shown in Table 4.

[0113] Table 4 PCR Procedure

[0114]

[0115] (6) Electrophoresis detection of PCR products: Take an appropriate amount of PCR products and load them onto a 2% to 3% agarose gel for electrophoresis to detect whether the amplification is successful.

[0116] Capillary electrophoresis detection, results as follows Figure 1 As shown:

[0117] (1) The PCR amplification products were centrifuged at 3500 rpm for 2 min using a benchtop or plate centrifuge.

[0118] (2) Prepare a mixture of HiDi and Liz500. After mixing the mixture with a vortex mixer for 10 seconds, centrifuge it with a microcentrifuge for 10 seconds and dispense 9 μL into a 96-well plate. Add 1 μL of PCR product supernatant (stock solution or dilution) to each well.

[0119] (3) Cover with a rubber pad and centrifuge instantly using a benchtop or plate centrifuge;

[0120] (4) 95℃, denature for 5 min, then immediately ice bath for 3 min;

[0121] (5) Perform capillary electrophoresis using a gene analyzer. Select the "Fragment" electrophoresis method. Data analysis is performed using analytical software (Applied Biosystems).

[0122] VI. Results Analysis

[0123] Table 5

[0124]

[0125] Table 6

[0126]

[0127] Genotyping results show:

[0128] For the SNP (rs334299237) locus, 236 pigs had the TT genotype, and 175 pigs had the TC genotype; for the SNP (rs318687930) locus, 246 pigs had the GG genotype, and 186 pigs had the GA genotype; for the SNP (rs325934988) locus, 263 pigs had the GG genotype, and 145 pigs had the GA genotype.

[0129] Table 5 shows the allele and gene frequencies of the CREBRF gene in the pig herd: For the SNP (rs334299237), the TT genotype frequency was 0.4155, and the TC genotype frequency was 0.3081. The TT genotype frequency was higher than the TC allele, indicating that the TT allele was the dominant gene. For the SNP (rs318687930), the GG genotype frequency was 0.4331, and the GA genotype frequency was 0.3275. The GG genotype frequency was higher than the GA allele, indicating that the GG allele was the dominant gene. For the SNP (rs325934988), the GG genotype frequency was 0.4630, and the GA genotype frequency was 0.2553. The GG genotype frequency was higher than the GA allele, indicating that the GG allele was the dominant gene.

[0130] Table 6 shows that: at the SNP (rs334299237) locus, there were 236 TT genotypes and 175 TC genotypes in 590 Large White pigs. The CREBRF showed a highly significant difference between the total number of piglets born in primiparous litters and the total number of piglets born after multiple births, with the TC genotype having a higher CREBRF than the TT genotype. At the SNP (rs318687930) locus, there were 246 GG genotypes and 186 GA genotypes in 590 Large White pigs. The CREBRF showed a highly significant difference between the total number of piglets born in primiparous litters and the total number of piglets born after multiple births, with the GA genotype having a higher CREBRF than the GG genotype. At the SNP (rs325934988) locus, there were 263 GG genotypes and 145 GA genotypes in 590 Large White pigs. The CREBRF showed a highly significant difference between the total number of piglets born in primiparous litters and the GA genotype having a higher CREBRF than the GG genotype.

[0131] Association analysis between genotype and total litter size phenotype was performed using a linear model in PLINK software, with field effect as a covariate for correction. The results are shown in Table 6. The SNP (rs334299237) locus significantly affected the total litter size in primiparous pigs. The TC genotype had a significantly higher total litter size in primiparous pigs than the TT genotype. Therefore, in actual pig breeding, TC genotype pigs have a higher total litter size in primiparous pigs. The SNP (rs318687930) locus significantly affected the total litter size in pigs. The GA genotype had significantly higher total litter size in both primiparous and multiparous pigs than the GG genotype. Therefore, in actual pig breeding, GA genotype pigs have a higher total litter size in total litter size. The SNP (rs325934988) locus significantly affected the total litter size in primiparous pigs. The GA genotype had a significantly higher total litter size in primiparous pigs than the GG genotype. Therefore, in actual pig breeding, GA genotype pigs have a higher total litter size in primiparous pigs.

[0132] In summary, determining the nucleotide sequence at the SNP (rs334299237) site of the CREBRF gene in pigs can help identify whether an individual has the TC or TT genotype, thus aiding in the identification of total litter size at primiparity: the TC genotype has a significantly higher total litter size at primiparity than the TT genotype. The TC genotype is a heterozygous combination of T and C at nucleotide position 51160002 on pig chromosome 16; the TT genotype is a homozygous combination of T at nucleotide position 51160002 on pig chromosome 16.

[0133] The nucleotide sequence at the SNP (rs318687930) site of the CREBRF gene in pigs can be used to determine whether an individual pig has the GA or GG genotype, thus aiding in the identification of primiparous total litter size and multiparous total litter size: the GA genotype shows significantly higher primiparous and multiparous total litter size than the GG genotype. The GA genotype is a heterozygous combination of G and A at nucleotide position 51174074 on pig chromosome 16; the GG genotype is a homozygous combination of G at nucleotide position 51174074 on pig chromosome 16.

[0134] The nucleotide sequence at the SNP (rs325934988) site of the CREBRF gene in pigs can be used to determine whether an individual pig has the GA or GG genotype, thus aiding in the identification of total litter size at primiparity: the GA genotype results in a significantly higher total litter size at primiparity than the GG genotype. The GA genotype is characterized by a heterozygous G / A sequence at position 51225561 on chromosome 16; the GG genotype is characterized by a homozygous G sequence at position 51225561 on chromosome 16.

[0135] Finally, it should be noted that the above embodiments are only used to illustrate the technical solutions of the present invention, rather than to limit it. Although the present invention has been described in detail with reference to the above embodiments, those skilled in the art should understand that they can still modify the technical solutions described in the above embodiments, or replace some or all of the technical features therein with equivalents. However, these modifications or replacements do not cause the essence of the corresponding technical solutions to deviate from the scope of the technical solutions of the embodiments of the present invention.

Claims

1. The application of substances that detect SNP sites in the preparation of products for predicting the total litter size trait in primiparous pigs, wherein the SNP sites include: One or more of the SNP loci rs334299237 and SNP loci rs325934988; The substance is a primer and / or a probe; The pig in question is a Large White pig.

2. A method for predicting the total number of piglets in a pig's first litter, characterized in that, This includes predicting sows that meet at least one of (i) to (ii) as having a high total litter size trait: (i) The genotype of SNP locus rs334299237 is TC. (ii) The genotype of SNP locus rs325934988 is GA; The pig in question is a Large White pig.

3. A method for screening sows with a high initial litter size trait, characterized in that, This includes screening sows with a high initial total litter size trait predicted by the method described in claim 2; The sow in question is a Large White pig.

4. A breeding method for sows with a high initial litter size trait, characterized in that, This includes acquiring at least one of the following genotypes in sows: (i) The genotype of SNP locus rs334299237 is TC. (ii) The genotype of SNP locus rs325934988 is GA; The sow in question is a Large White pig.

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