Analysis of ESR1 Gene Polymorphism and Lambing Association between Cele Black Sheep and Hetian Sheep

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The purpose of this study is to detect the ESR1 gene in Cele Black sheep and Hetian sheep populations, so as to lay a foundation for further improving the litter size of the population and establishing a core population with high fecundity. Jugular blood DNA was extracted from healthy Cele Black sheep and Hetian sheep, and two polymorphic loci (rs399356740 and rs590747651) of ESR1 gene in two sheep breeds were genotyped by Polymerase Chain Reaction (PCR) and Sanger sequencing, and the genetic parameters and lambing number were analyzed. The results of sequencing showed that there were three genotypes of AA, AT and TT at rs399356740 and rs590747651 loci in Cele Black sheep and Hetian sheep, and all of them showed moderate polymorphism. The dominant allele of rs590747651 locus in Cele Black sheep is T and the dominant allele of rs590747651 locus in Hetian sheep is A, which may be the reason for the difference in fecundity between the two sheep populations. The mutation site of rs590747651 was in Hardy-Weinberg equilibrium in Hetian sheep population, and the results of association analysis showed that the lambing number of AA and TT type at rs590747651 locus was significantly higher than that of AT type in Hetian sheep population, and the dominant genotype was AA type, which indicated that rs590747651 mutation could be considered as a molecular assistant marker for lambing trait in Hetian sheep. However, due to the small sample size, the rs590747651 locus had nothing to do with the litter size of Cele Black sheep. This study can be used as a potential candidate genetic marker for lambing performance in sheep breeding, and provide reference data for the study of lambing performance of sheep. ESR1 gene polymorphism lamb Cele Black sheep Hetian sheep Figures Figure 1 Figure 2 1 Introduction Litter size is one of the most important reproductive traits of sheep, and it has always been considered as a key index affecting reproductive performance and productivity. Low litter size is the biggest bottleneck restricting the productivity of ewes, and its heritability is low, so it is difficult for conventional breeding techniques to rapidly improve the progress of breeding. At present, the most economical and effective way to improve fecundity is to explore relevant important candidate genes and carry out marker-assisted selection of candidate loci, because it can greatly improve the selection efficiency of these low heritability traits by affecting selection time, intensity and accuracy, and is suitable for improving litter size traits. At present, many studies have reported that estrogen receptor (ESR), growth differentiation factor 9 (GDF9), bone morphogenetic protein 15 (BMP15) and bone morphogenetic proteblackin receptor 1B (BMPR1B) play an important role in the regulation of lambing traits in sheep [1]. There are also different lambing numbers within and among sheep breeds. Cele Black sheep and Hetian sheep are two local breeds unique to Xinjiang, China, with the characteristics of high fecundity and low fecundity, respectively. Cele Black sheep has the characteristics of high reproduction rate, annual estrus, the average lambing rate is 215.5%; Hetian sheep has the disadvantage of low reproduction rate, the average lambing rate is 102.5%. There is a significant difference in fecundity between them, which provides a suitable animal model marker for the identification and utilization of main reproductive genes in sheep [2–4]. Estrogen receptor 1 ( ESR1 ) is considered to be the main estrogen receptor mediating the regulation of GnRH secretion by estradiol, In the early years, it was proposed as the main gene of reproduction, which, when combined with estrogen, plays an important role in the growth and development of follicles in the embryo, mammary gland and female reproductive cycle [5–6]. S.C.Hewitt (2003) found that ESR knockout mice do not ovulate, luteinizing hormone (LH) regulation is disordered, and the uterus is not sensitive to estrogen, which indicates that ESR plays an important role in reproduction [7]. In recent years, many studies have focused on the analysis of the association between ESR gene polymorphism and litter traits.In 1996, the polymorphism of ESR PvuII restriction site was found for the first time in large White pigs, which was significantly correlated with litter size [8]. Liu Qingqing et al (2023) found that the rs399356740 locus of ESR1 gene can provide a valuable reference for the selection of litter size traits in small-tailed Han sheep, and the missense mutation of ESR1 may reduce its ability to bind to NCOA1, resulting in a decrease in fecundity of small-tailed Han sheep [9]. In this experiment , PCR amplification and direct sequencing were used to analyze the genetic polymorphism of two single nucleotide polymorphism (SNP) loci of ESR1 gene in Cele Black sheep and Hetian sheep, and to explore the relationship between the identified SNP loci and lambing traits, in order to provide some effective genetic markers for molecular breeding of sheep. 2 Article types Test sample collection n this experiment, 103 Cele Black sheep and 105 Hetian sheep, a total of 208 2–4 year old healthy ewes were selected as test samples.The blood samples of Cele Black sheep were collected from Kunlun Luyuan Sheep Housekeeping Animal Farm in Cele County, Hetian area, and the blood samples of Hetian Sheep were collected from Hetian Sheep breeding Farm in Luopu County. All the blood samples were collected from jugular vein (10ml/pcs) by negative pressure anticoagulant blood collection and temporarily preserved in dry ice, After taking it back to the laboratory, store it in the refrigerator at-80℃ for use. Main reagents and instruments 2× Easy Taq® PCR SuperMix (+ Dye) and DNA Marker were purchased from Beijing TransGen Biotechnology Co., Ltd., agar powder, EDTA and TAE buffers were purchased from Xinjiang Taklan Biotechnology Co., Ltd., gradient PCR instrument, Bio-Rad PAC 3000 electrophoresis instrument and ChemiDoc chemiluminescence imaging analysis system were purchased from Bio-Rad (USA) company, the electrophoresis tank was purchased from Beijing Liuyi Instrument Factory, and the micropipette was purchased from Eppendorf. Extraction of DNA from blood Sheep genomic DNA was extracted using the Tiangen blood/cell/tissue genomic DNA extraction kit. The concentration of DNA samples was detected by DeNovix DS-11 ultra-microvolume spectrophotometer (DeNovix,USA), and then the DNA concentration and purity were detected by 2% agarose gel electrophoresis. Primer design and synthesis According to the sequence of sheep ESR1 gene in NCBI database (accession number: NC_056061.1) and two polymorphic loci (rs39935740 and rs590747651) found in Ensembl database, the specific primers were designed by Primer5 software. Two pairs of primers were designed, rs39935740 primer sequence 5'-GTAACAATGTACCTCAGCCTGGA and 3'-GGCAGATACCCACACAGAGATAT (Tm = 54℃), rs590747651 primer sequence 5'-AGGCAACCCAATCTCTGTGAC and 3'-CTTCCCTGATCCCACAACCA (Tm = 57℃), the length of PCR product was 341bp and 384bp respectively, the primer was synthesized by Sangon Biotech (Shanghai) Co., Ltd. PCR amplification and sequencing DNA of Cele Black sheep and Hetian sheep was used as PCR template to amplify ESR1 gene. 25 µL PCR reaction system: 12.5 µL 2 × Easy Taq ®PCR SuperMix (+ Dye), 9.5 µL ddH2O, 1 µL DNA, 1 µL upstream primer and 1 µL downstream primer (10nmol/L). PCR reaction program: predenaturation at 95℃ for 3 min, denaturation at 95℃ for 15s, annealing at 54℃ for 20s, extension at 72℃ for 30s, a total of 31 cycles, extension at 72℃ for 6 min and preservation at 4℃. The PCR products were detected by 2% agarose gel electrophoresis, and the electrophoresis condition was 150V, 130mA and 25-40min. The successfully amplified PCR products were sent to Sangong Biotech (Shanghai) Co., Ltd for sequencing, and the sequencing peak map was analyzed by SnapGene software. Statistical analysis In this experiment, the typing data obtained by PCR amplification and sequencing were analyzed by Excel software, statistical analysis of genetic parameters of two polymorphic loci (rs39935740 and rs590747651) of ESR1 gene. Genotype frequency, allele frequency, heterozygosity (He), effective number of alleles (Ne), p value and polymorphism information content (PIC) were calculated by formula and Excel software. P value was calculated by SPSS 25.0, Chi-square test p > 0.05 showed that the locus was in Hardy-Weinberg equilibrium. The least square method of SPSS 25.0 was used to analyze the correlation between different genotypes and litter size in different sheep populations. The calculation formula is as follows: He = 1- \(\:\sum\:_{\mathbf{i}=1}^{\mathbf{n}}{\mathbf{P}\mathbf{i}}^{2}\) Ne = 1/ \(\:\sum\:_{\mathbf{i}=1}^{\mathbf{n}}{\mathbf{P}\mathbf{i}}^{2}\) $$\:\mathbf{P}\mathbf{I}\mathbf{C}\hspace{0.17em}=\hspace{0.17em}1-\sum\:_{\mathbf{i}=1}^{\mathbf{n}}{\mathbf{P}\mathbf{i}}^{2}-\sum\:_{\mathbf{i}=1}^{\mathbf{n}-1}\sum\:_{\mathbf{j}=\mathbf{i}+1}^{\mathbf{n}}{2\mathbf{P}\mathbf{i}}^{2}{\mathbf{P}\mathbf{j}}^{2}$$ i is the i allele, j is the j allele, Pi and Pj are the i and j allele frequencies, and n is the number of multiple alleles. 3 Results Analysis of ESR1 gene sequencing and SNP polymorphism According to the designed primers, using the DNA of Cele Black sheep (n = 103) and Hetian sheep (n = 105) as templates to amplificate, the specific bands of rs399356740 and rs590747651 loci were obtained from left to right, and the fragments were 341bp and 384bp(Fig. 1 ), which were consistent with the expected fragment size. Two SNP loci of ESR1 gene were amplified, and the sequencing peak maps were compared by SnapGene software, the genotypes of two SNP loci were obtained, both of which were intron mutations. When there are two obvious peaks at the same site, the sample is heterozygote, and when there is only one peak at the same site, the sample is homozygous. The results of PCR sequencing of Cele Black sheep (n = 103) and Hetian sheep (n = 105) are shown in Fig. 2 , and the two candidate loci of ESR1 gene have polymorphisms. There are three genotypes of rs399356740 locus and rs590747651 locus in Cele Black sheep and Hetian sheep, which are AA, AT and TT, respectively. Genetic diversity index of ESR1 gene in Cele Black sheep The differences of genotype frequency and allele frequency of different loci of ESR1 gene in single lamb and multi-lamb of Cele Black sheep were shown in Table 1 , in which the genotype frequency and allele frequency of rs590747651 locus were statistically significant in single lamb and multi-lamb. As far as rs590747651 locus is concerned, AT is the dominant genotype, and the dominant allele in multi-lambs is A. Table 2 showed that the rs590747651 locus of ESR1 gene was moderately polymorphic in both single lambs and multiple lambs of Black sheep (0.25 ≤ PIC < 0.5). Genetic diversity index of ESR1 gene in Hetian sheep The differences of genotype frequency and allele frequency of different loci of ESR1 gene in single lamb and multi-lamb of Hetian sheep were shown in Table 3 , in which the genotype frequency and allele frequency of rs399356740 locus were statistically significant in single lamb and multi-lamb. In terms of rs399356740 loci, AT was the dominant genotype, T was the dominant allele in single lamb breeds, AA was the dominant genotype and A was the dominant allele in multi-lamb breeds. Table 4 showed that the rs399356740 locus of ESR1 gene was moderately polymorphic in both single lambs and multiple lambs of Hetian sheep (0.25 ≤ PIC < 0.5). Table 1 Gene frequencies and genotype frequencies of different loci of ESR1 gene in single and multiple lambs in Cele Black sheep. Locus Genotype Genotype frequency in uniparous sheep(sample size) Genotype frequency in multiparous sheep(sample size) P value Allele Allele frequency in uniparous sheep Allele frequency in multiparous sheep P value rs399356740 AA 0.27(13) 0.15(8) 0.06 A 0.43(21) 0.31(17) 0.23 AT 0.69(34) 0.85(46) T 0.57(28) 0.69(37) TT 0.04(2) 0.00(0) rs590747651 AA 0.02(1) 0.00(0) 0.04 A 0.37(18) 0.59(32) 0.02 AT 0.92(45) 1.00(54) T 0.63(31) 0.41(22) TT 0.06(3) 0.00(0) P < 0.05 indicates the difference was significant. Table 2 Polymorphic information content, heterozygosity and effective alleles of different loci of ESR1 gene in Cele Black sheep. Locus He in uniparous sheep He in multiparous sheep PIC in uniparous sheep PIC in multiparous sheep Ne in uniparous sheep Ne in multiparous sheep rs399356740 0.49 0.43 0.37 0.34 1.96 1.75 rs590747651 0.47 0.48 0.36 0.37 1.87 1.94 Table 3 Gene frequencies and genotype frequencies of different loci of ESR1 gene in single and multiple lambs in Hetian sheep. Locus Genotype Genotype frequency in uniparous sheep(sample size) Genotype frequency in multiparous sheep(sample size) P value Allele Allele frequency in uniparous sheep Allele frequency in multiparous sheep P value rs399356740 AA 0.34(21) 0.56(24) 0.04 A 0.37(23) 0.60(26) 0.02 AT 0.61(38) 0.44(19) T 0.63(39) 0.40(17) TT 0.05(3) 0.00(0) rs590747651 AA 0.21(13) 0.39(17) 0.05 A 0.66(41) 0.65(28) 0.27 AT 0.50(31) 0.28(12) T 0.34(21) 0.35(15) TT 0.29(18) 0.33(14) P < 0.05 indicates the difference was significant. Table 4 Polymorphic information content, heterozygosity and effective alleles of ESR1 gene at different loci of ESR1 gene in Hetian sheep. Locus He in uniparous sheep He in multiparous sheep PIC in uniparous sheep PIC in multiparous sheep Ne in uniparous sheep Ne in multiparous sheep rs399356740 0.47 0.48 0.36 0.36 1.87 1.92 rs590747651 0.45 0.46 0.35 0.35 1.81 1.83 Population genetic analysis of different SNPs of ESR1 gene in different sheep breeds The loci of rs399356740 and rs590747651 of ESR1 gene were moderately polymorphic in Cele Black sheep and Hetian sheep (0.25 < PIC < 0.5). Chi-square test showed that the ESR1 gene rs399356740 locus was in Hardy-Weinberg disequilibrium in Cele Black sheep and Hetian sheep breeds, and the rs590747651 locus in ESR1 gene was in Hardy-Weinberg disequilibrium in Cele Black sheep and was in Hardy-Weinberg equilibrium in Hetian sheep. Cele Black sheep is a high yield sheep, but Hetian sheep is not. in the two loci of ESR1 gene rs399356740 and rs590747651, the dominant allele of rs399356740 in the two sheep breeds is T (Table 5 ), but the dominant allele of rs590747651 in Cele Black sheep is T, and the dominant allele in Hetian sheep is A, so it is speculated that this may be the reason for the difference in fecundity between the two sheep populations. Table 5 Population genetic analysis of different loci of ESR1 gene in different sheep breeds. Locus Variety Genotype frequency(sample size) Allele Frequency Polymorphism information content(PIC) Heterozy gosity (He) Effective number of allele(Ne) P value AA AT TT A T rs399356740 Cele Black sheep 0.20(21) 0.78(80) 0.02(2) 0.37 (38) 0.63 (65) 0.36 0.47 1.87 0.00 Hetian sheep 0.43(45) 0.54(57) 0.03(3) 0.47 (49) 0.53 (56) 0.37 0.50 1.99 0.00 rs590747651 Cele Black sheep 0.01(1) 0.96(99) 0.03(3) 0.49 (50) 0.51 (53) 0.37 0.50 1.99 0.00 Hetian sheep 0.29(30) 0.41(43) 0.30(32) 0.66 (69) 0.34 (36) 0.35 0.45 1.81 0.06 P < 0.05 indicates the difference was significant. Analysis of the correlation between SNP locus of ESR1 gene and litter size Table 6 showed that there was no significant correlation between rs399356740 genotypes of ESR1 gene and litter size in Cele Black sheep and Hetian sheep, and there was no significant correlation between genotypes of ESR1 gene rs590747651 locus and litter size of Cele Black sheep. The lambing number of AA and TT genotypes of ESR1 gene rs590747651 mutation in Hetian sheep was significantly higher than that of AT type, and there was no significant difference between AA and TT genotypes. Table 6 Correlation Analysis of SNP Locus of ESR1 Gene and litter size of Cele Black sheep and Hetian sheep. Variety SNP Genotype Number of lambs Cele Black sheep rs399356740 AA 1.48 ± 0.50 AT 1.50 ± 0.50 TT 1.00 ± 0.00 rs590747651 AA 1.00 ± 0.00 AT 1.55 ± 0.50 TT 1.00 ± 0.00 Hetian sheep rs399356740 AA 1.53 ± 0.50 AT 1.33 ± 0.48 TT 1.00 ± 0.00 rs590747651 AA 1.57 ± 0.50 a AT 1.28 ± 0.45 b Note: when comparing different genotypes, the difference is significant when there is any letter difference after the data (P < 0.05). 4 Discussion Cele Black sheep and Hetian sheep are two local sheep breeds unique to Xinjiang, China, and their reproductive performance is quite different, among which the lambing rate of Hetian sheep is low, which seriously hinders the economic development of the local sheep industry. However, Hetian sheep, as a characteristic sheep germplasm resource in Xinjiang, has formed the germplasm characteristics of drought and heat tolerance and strong adaptability under the unique natural ecological conditions of Hetian region, Ewes can be in estrus all the year round, and the lambing rate of Cele Black sheep is high. Therefore, the breeding of Cele Black sheep and Hetian sheep plays an important role in promoting the overall sheep breeding and economic development in Xinjiang [10]. Bai Shu et al (2013) found that ESR1 was expressed in uterine lumen epithelium, glandular epithelium, matrix, vascular endothelium and smooth muscle cell nucleus in Jining grey goats, which confirmed that ESR1 was involved in regulating the proliferation and differentiation of endometrium and myometrium [11].After knocking out the ESR gene of female mice, ESR-deficient female mice appeared luteinizing hormone regulation disorder and ovary anovulation, which affected the fertility of mice [7]. Overexpression of ESR1 may lead to follicular atresia [12]. Generally speaking, ESR1 may have a certain effect on reproductive performance, and ESR1 gene, as one of the subtypes of ESR, plays an important role in lambing performance of mammals. In early experiments, it has been confirmed that ESR1 in endometrium is necessary to identify pregnancy and maternal development in pigs [13, 14]. Rempel et al found that the SNP loci of ESR1 and ESR2 were significantly correlated with litter size. The high yield genotype of ESR gene Pvu II restriction site of Baoshan pig was AB type, and the B allele had a positive effect on the litter size of Baoshan sows. Shun Wu et al found that the TT genotype of ESR g.C1665T locus and the AA genotype of g.A1755G locus could significantly increase the total litter size of the first birth of American large White pigs [15–17]. There are many studies of ESR1 gene on the reproductive performance of other mammals, such as duck [18], quail [19] and fish [20]. These studies show that ESR1 gene plays a key role in estrus ovulation and lambing. The purpose of this study is to identify ESR1 gene as a potential molecular marker and further study the relationship between MAS and lambing, so as to apply MAS to sheep breeding. It was reported that there was a significant correlation between the ESR1 locus rs399356740 locus and the number of Small-Tailed Han lambs, and the number of lambs in AA ewes was significantly higher than that in TT ewes, but there was no significant correlation between rs590747651 locus and the number of lambs in different small-tailed Han sheep, that is, there was more and more evidence that mutations in ESR1 gene locus were related to animal reproduction [21]. The current study aimed to check the association of ESR1 rs399356740 and rs590747651 with the litter size in the Cele Black sheep and Hetian sheep. We found that the rs399356740 and rs590747651 loci of ESR1 gene in Cele Black sheep and Hetian sheep were moderately polymorphic. Except that the rs590747651 locus was in Hardy-Weinberg equilibrium in Hetian sheep, none of them showed Hardy-Weinberg equilibrium, which may be due to site imbalance caused by natural selection or artificial intervention, or due to the small number of sheep involved in this study. Our study shows that the rs399356740 and Cele Black sheep of ESR1 gene have nothing to do with the litter size of Hetian sheep, and rs590747651 has nothing to do with the litter size of Cele Black sheep, which may be caused by the lack of high selection in the breeding process, but it can not be ruled out that it is due to the small sample size. However, there was a significant correlation between rs590747651 and lambing number of Hetian sheep. The lambing number of AA and TT type ewes was significantly higher than that of AT type ewes. Therefore, in the process of Hetian sheep breeding, homozygous individuals of AA and TT can be retained to improve the fecundity of Hetian sheep. This study provided a preliminary reference for marker-assisted selection of litter size in Hetian sheep, but in view of the shortage of samples, the TT genotypes of the detected rs399356740 loci in Cele Black sheep and Hetian sheep, and the number of AA and TT genotypes of rs590747651 in Cele Black sheep were insufficient, it is necessary to expand the sample size and improve the accuracy of the experiment. 5 Conclusions This study summarized the association between ESR1 gene rs590747651 and rs399356740 and lambing number of Cele black sheep and Hetian sheep, and found that the variation of rs399356740 was related to the litter size of Hetian sheep. Statistical analysis showed that the litter size of ESR1 rs399356740 homozygous genotype was significantly higher than that of heterozygous genotype. Abbreviations Item Unit PCR Polymerase Chain Reaction ESR Estrogen Receptor GDF9 Growth Differentiation Factor 9 BMP15 Bone Morphogenetic Protein 15 BMPR1B Bone Morphogenetic Protein Receptor 1B ESR1 Estrogen Receptor 1 GnRH Gonadotropin-Releasing Hormone LH Luteinizing Hormone SNP Single Nucleotide Polymorphism Declarations 7.1 Ethics approval and consent to participate For experimental animals, all protocols were performed in accordance with the “Guide to Animal Experimentation” and approved by the Use Committee under the norms of the Ethics Committee of Tarim University of Science and Technology (SYXK 2020-009). 7.2 Consent for publication Not applicable. This manuscript does not contain personal data from any individual. All authors have reviewed the final version of the manuscript and consent to its publication in BMC Genomics. 7.3 Availability of data and materials The datasets generated and analyzed during this study are included in this published article and its supplementary files. The raw sequencing data are available from the corresponding author upon reasonable request. Biological materials: Blood samples from Cele Black sheep and Hetian sheep are preserved at the Key Laboratory of Utilization of Surrounding Tarim Livestock and Grass Resources (Tarim University) and may be requested for non-commercial research purposes. Public data resources: Reference sequences for the ESR1 gene (NCBI Accession: NC_056061.1) and SNP loci (rs399356740, rs590747651) were obtained from the NCBI (https://www.ncbi.nlm.nih.gov/) and Ensembl (https://www.ensembl.org/) databases. Primers: All primer sequences used for PCR amplification are provided in the Methods section. 7.4 Competing interests. The authors declare that this research was conducted in the absence of any conflicts of interest. 7.5 Funding. The study was made possible thanks to funding from the Xinjiang Production and Construction Corps (Grant/Award Numbers: 2022CB00104) and the President's Fund Project of Tarim University (Grant/Award Numbers: TDZKCX202401). 7.6 Author Contributions. Q.H., G.R conceived the study, conducted data analysis, and prepared the numbers and tables. Q.H., G.R. and W.L. collected the samples, L.Z, and H.S. conducted genetic diversity analyses F.X. supervised the study. Q.H. This manuscript has been prepared. All the authors have read and agreed to the published version of the manuscript. References Yue C, Bai WL, Zheng YY, Hui TY, Sun JM, Guo D, Guo SL. Correlation. Analysis of candidate gene SNP for high-yield in Liaoning cashmere goats with litter size and cashmere performance. 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14:35:36","extension":"html","order_by":9,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":82009,"visible":true,"origin":"","legend":"","description":"","filename":"earlyproof.html","url":"https://assets-eu.researchsquare.com/files/rs-7326024/v1/6273d8344446c37d70d0c968.html"},{"id":92518298,"identity":"c8d895c1-27dd-45ee-975a-ca0f828c5190","added_by":"auto","created_at":"2025-09-30 14:35:35","extension":"jpeg","order_by":1,"title":"Figure 1","display":"","copyAsset":false,"role":"figure","size":17522,"visible":true,"origin":"","legend":"\u003cp\u003eAmplification of \u003cem\u003eESR1\u003c/em\u003e gene polymorphisms. M, DNA Marker; 1~3, rs399356740 site amplification product; 4~6, rs590747651 site amplification product.\u003c/p\u003e","description":"","filename":"floatimage1.jpeg","url":"https://assets-eu.researchsquare.com/files/rs-7326024/v1/4da9c97309377006fe31fb9f.jpeg"},{"id":92519538,"identity":"00427b9d-9a04-4c89-a82d-efc2c29b6a46","added_by":"auto","created_at":"2025-09-30 14:43:35","extension":"jpeg","order_by":2,"title":"Figure 2","display":"","copyAsset":false,"role":"figure","size":232909,"visible":true,"origin":"","legend":"\u003cp\u003ePeak plot of \u003cem\u003eESR1\u003c/em\u003e gene SNP locus by PCR. A: PCR sequencing peak of \u003cem\u003eESR1\u003c/em\u003e gene rs399356740 locus; B: PCR sequencing peak of \u003cem\u003eESR1\u003c/em\u003e gene rs590747651 locus.\u003c/p\u003e\n\u003cp\u003eNote: AA and TT is homozygous, AT is heterozygous.\u003c/p\u003e","description":"","filename":"floatimage2.jpeg","url":"https://assets-eu.researchsquare.com/files/rs-7326024/v1/0ce2b58ab969349f57f01a35.jpeg"},{"id":96708128,"identity":"86328e7e-b754-4512-8e32-6938ff266332","added_by":"auto","created_at":"2025-11-25 09:57:51","extension":"pdf","order_by":0,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":1233646,"visible":true,"origin":"","legend":"","description":"","filename":"manuscript.pdf","url":"https://assets-eu.researchsquare.com/files/rs-7326024/v1/fb3ec071-f508-4352-b609-8fa5fdde11e0.pdf"},{"id":92518307,"identity":"3dd000f8-53b2-4b6e-bae5-d56d67e5f8b7","added_by":"auto","created_at":"2025-09-30 14:35:35","extension":"jpg","order_by":0,"title":"","display":"","copyAsset":false,"role":"supplement","size":626452,"visible":true,"origin":"","legend":"","description":"","filename":"SupplementaryFile1UncroppedGelFig1.tif.jpg","url":"https://assets-eu.researchsquare.com/files/rs-7326024/v1/980e1012d77e42d2fcb19210.jpg"}],"financialInterests":"No competing interests reported.","formattedTitle":"\u003cp\u003eAnalysis of \u003cem\u003eESR1\u003c/em\u003e Gene Polymorphism and Lambing Association between Cele Black Sheep and Hetian Sheep\u003c/p\u003e","fulltext":[{"header":"1 Introduction","content":"\u003cp\u003eLitter size is one of the most important reproductive traits of sheep, and it has always been considered as a key index affecting reproductive performance and productivity. Low litter size is the biggest bottleneck restricting the productivity of ewes, and its heritability is low, so it is difficult for conventional breeding techniques to rapidly improve the progress of breeding. At present, the most economical and effective way to improve fecundity is to explore relevant important candidate genes and carry out marker-assisted selection of candidate loci, because it can greatly improve the selection efficiency of these low heritability traits by affecting selection time, intensity and accuracy, and is suitable for improving litter size traits. At present, many studies have reported that estrogen receptor (ESR), growth differentiation factor 9 (GDF9), bone morphogenetic protein 15 (BMP15) and bone morphogenetic proteblackin receptor 1B (BMPR1B) play an important role in the regulation of lambing traits in sheep [1]. There are also different lambing numbers within and among sheep breeds. Cele Black sheep and Hetian sheep are two local breeds unique to Xinjiang, China, with the characteristics of high fecundity and low fecundity, respectively. Cele Black sheep has the characteristics of high reproduction rate, annual estrus, the average lambing rate is 215.5%; Hetian sheep has the disadvantage of low reproduction rate, the average lambing rate is 102.5%. There is a significant difference in fecundity between them, which provides a suitable animal model marker for the identification and utilization of main reproductive genes in sheep [2\u0026ndash;4].\u003c/p\u003e\u003cp\u003eEstrogen receptor 1 (\u003cem\u003eESR1\u003c/em\u003e) is considered to be the main estrogen receptor mediating the regulation of GnRH secretion by estradiol, In the early years, it was proposed as the main gene of reproduction, which, when combined with estrogen, plays an important role in the growth and development of follicles in the embryo, mammary gland and female reproductive cycle [5\u0026ndash;6]. S.C.Hewitt (2003) found that ESR knockout mice do not ovulate, luteinizing hormone (LH) regulation is disordered, and the uterus is not sensitive to estrogen, which indicates that ESR plays an important role in reproduction [7]. In recent years, many studies have focused on the analysis of the association between ESR gene polymorphism and litter traits.In 1996, the polymorphism of ESR PvuII restriction site was found for the first time in large White pigs, which was significantly correlated with litter size [8]. Liu Qingqing et al (2023) found that the rs399356740 locus of \u003cem\u003eESR1\u003c/em\u003e gene can provide a valuable reference for the selection of litter size traits in small-tailed Han sheep, and the missense mutation of \u003cem\u003eESR1\u003c/em\u003e may reduce its ability to bind to NCOA1, resulting in a decrease in fecundity of small-tailed Han sheep [9].\u003c/p\u003e\u003cp\u003eIn this \u003cb\u003eexperiment\u003c/b\u003e, PCR amplification and direct sequencing were used to analyze the genetic polymorphism of two single nucleotide polymorphism (SNP) loci of \u003cem\u003eESR1\u003c/em\u003e gene in Cele Black sheep and Hetian sheep, and to explore the relationship between the identified SNP loci and lambing traits, in order to provide some effective genetic markers for molecular breeding of sheep.\u003c/p\u003e"},{"header":"2 Article types","content":"\u003cp\u003eTest sample collection\u003c/p\u003e\u003cp\u003en this experiment, 103 Cele Black sheep and 105 Hetian sheep, a total of 208 2\u0026ndash;4 year old healthy ewes were selected as test samples.The blood samples of Cele Black sheep were collected from Kunlun Luyuan Sheep Housekeeping Animal Farm in Cele County, Hetian area, and the blood samples of Hetian Sheep were collected from Hetian Sheep breeding Farm in Luopu County. All the blood samples were collected from jugular vein (10ml/pcs) by negative pressure anticoagulant blood collection and temporarily preserved in dry ice, After taking it back to the laboratory, store it in the refrigerator at-80℃ for use.\u003c/p\u003e\u003cp\u003eMain reagents and instruments\u003c/p\u003e\u003cp\u003e2\u0026times; Easy Taq\u0026reg; PCR SuperMix (+\u0026thinsp;Dye) and DNA Marker were purchased from Beijing TransGen Biotechnology Co., Ltd., agar powder, EDTA and TAE buffers were purchased from Xinjiang Taklan Biotechnology Co., Ltd., gradient PCR instrument, Bio-Rad PAC 3000 electrophoresis instrument and ChemiDoc chemiluminescence imaging analysis system were purchased from Bio-Rad (USA) company, the electrophoresis tank was purchased from Beijing Liuyi Instrument Factory, and the micropipette was purchased from Eppendorf.\u003c/p\u003e\u003cp\u003eExtraction of DNA from blood\u003c/p\u003e\u003cp\u003eSheep genomic DNA was extracted using the Tiangen blood/cell/tissue genomic DNA extraction kit. The concentration of DNA samples was detected by DeNovix DS-11 ultra-microvolume spectrophotometer (DeNovix,USA), and then the DNA concentration and purity were detected by 2% agarose gel electrophoresis.\u003c/p\u003e\u003cp\u003ePrimer design and synthesis\u003c/p\u003e\u003cp\u003eAccording to the sequence of sheep \u003cem\u003eESR1\u003c/em\u003e gene in NCBI database (accession number: NC_056061.1) and two polymorphic loci (rs39935740 and rs590747651) found in Ensembl database, the specific primers were designed by Primer5 software. Two pairs of primers were designed, rs39935740 primer sequence 5'-GTAACAATGTACCTCAGCCTGGA and 3'-GGCAGATACCCACACAGAGATAT (Tm\u0026thinsp;=\u0026thinsp;54℃), rs590747651 primer sequence 5'-AGGCAACCCAATCTCTGTGAC and 3'-CTTCCCTGATCCCACAACCA (Tm\u0026thinsp;=\u0026thinsp;57℃), the length of PCR product was 341bp and 384bp respectively, the primer was synthesized by Sangon Biotech (Shanghai) Co., Ltd.\u003c/p\u003e\u003cp\u003ePCR amplification and sequencing\u003c/p\u003e\u003cp\u003eDNA of Cele Black sheep and Hetian sheep was used as PCR template to amplify \u003cem\u003eESR1\u003c/em\u003e gene. 25 \u0026micro;L PCR reaction system: 12.5 \u0026micro;L 2 \u0026times; Easy Taq \u0026reg;PCR SuperMix (+\u0026thinsp;Dye), 9.5 \u0026micro;L ddH2O, 1 \u0026micro;L DNA, 1 \u0026micro;L upstream primer and 1 \u0026micro;L downstream primer (10nmol/L). PCR reaction program: predenaturation at 95℃ for 3 min, denaturation at 95℃ for 15s, annealing at 54℃ for 20s, extension at 72℃ for 30s, a total of 31 cycles, extension at 72℃ for 6 min and preservation at 4℃. The PCR products were detected by 2% agarose gel electrophoresis, and the electrophoresis condition was 150V, 130mA and 25-40min. The successfully amplified PCR products were sent to Sangong Biotech (Shanghai) Co., Ltd for sequencing, and the sequencing peak map was analyzed by SnapGene software.\u003c/p\u003e\u003cp\u003eStatistical analysis\u003c/p\u003e\u003cp\u003eIn this experiment, the typing data obtained by PCR amplification and sequencing were analyzed by Excel software, statistical analysis of genetic parameters of two polymorphic loci (rs39935740 and rs590747651) of \u003cem\u003eESR1\u003c/em\u003e gene. Genotype frequency, allele frequency, heterozygosity (He), effective number of alleles (Ne), p value and polymorphism information content (PIC) were calculated by formula and Excel software. P value was calculated by SPSS 25.0, Chi-square test p\u0026thinsp;\u0026gt;\u0026thinsp;0.05 showed that the locus was in Hardy-Weinberg equilibrium. The least square method of SPSS 25.0 was used to analyze the correlation between different genotypes and litter size in different sheep populations. The calculation formula is as follows:\u003c/p\u003e\u003cp\u003e\u003cb\u003eHe\u0026thinsp;=\u0026thinsp;1-\u003c/b\u003e\u003cspan class=\"InlineEquation\"\u003e\u003cspan class=\"mathinline\"\u003e\\(\\:\\sum\\:_{\\mathbf{i}=1}^{\\mathbf{n}}{\\mathbf{P}\\mathbf{i}}^{2}\\)\u003c/span\u003e\u003c/span\u003e\u003c/p\u003e\u003cp\u003e\u003cb\u003eNe\u0026thinsp;=\u0026thinsp;1/\u003c/b\u003e\u003cspan class=\"InlineEquation\"\u003e\u003cspan class=\"mathinline\"\u003e\\(\\:\\sum\\:_{\\mathbf{i}=1}^{\\mathbf{n}}{\\mathbf{P}\\mathbf{i}}^{2}\\)\u003c/span\u003e\u003c/span\u003e\u003cdiv id=\"Equa\" class=\"Equation\"\u003e\u003cdiv format=\"TEX\" class=\"mathdisplay\" id=\"FileID_Equa\" name=\"EquationSource\"\u003e\n$$\\:\\mathbf{P}\\mathbf{I}\\mathbf{C}\\hspace{0.17em}=\\hspace{0.17em}1-\\sum\\:_{\\mathbf{i}=1}^{\\mathbf{n}}{\\mathbf{P}\\mathbf{i}}^{2}-\\sum\\:_{\\mathbf{i}=1}^{\\mathbf{n}-1}\\sum\\:_{\\mathbf{j}=\\mathbf{i}+1}^{\\mathbf{n}}{2\\mathbf{P}\\mathbf{i}}^{2}{\\mathbf{P}\\mathbf{j}}^{2}$$\u003c/div\u003e\u003c/div\u003e\u003c/p\u003e\u003cp\u003ei is the i allele, j is the j allele, Pi and Pj are the i and j allele frequencies, and n is the number of multiple alleles.\u003c/p\u003e"},{"header":"3 Results","content":"\u003cp\u003eAnalysis of \u003cem\u003eESR1\u003c/em\u003e gene sequencing and SNP polymorphism\u003c/p\u003e\n\u003cp\u003eAccording to the designed primers, using the DNA of Cele Black sheep (n\u0026thinsp;=\u0026thinsp;103) and Hetian sheep (n\u0026thinsp;=\u0026thinsp;105) as templates to amplificate, the specific bands of rs399356740 and rs590747651 loci were obtained from left to right, and the fragments were 341bp and 384bp(Fig.\u0026nbsp;\u003cspan class=\"InternalRef\"\u003e1\u003c/span\u003e), which were consistent with the expected fragment size.\u003c/p\u003e\n\u003cp\u003eTwo SNP loci of \u003cem\u003eESR1\u003c/em\u003e gene were amplified, and the sequencing peak maps were compared by SnapGene software, the genotypes of two SNP loci were obtained, both of which were intron mutations. When there are two obvious peaks at the same site, the sample is heterozygote, and when there is only one peak at the same site, the sample is homozygous. The results of PCR sequencing of Cele Black sheep (n\u0026thinsp;=\u0026thinsp;103) and Hetian sheep (n\u0026thinsp;=\u0026thinsp;105) are shown in Fig.\u0026nbsp;\u003cspan class=\"InternalRef\"\u003e2\u003c/span\u003e, and the two candidate loci of \u003cem\u003eESR1\u003c/em\u003e gene have polymorphisms. There are three genotypes of rs399356740 locus and rs590747651 locus in Cele Black sheep and Hetian sheep, which are AA, AT and TT, respectively.\u003c/p\u003e\n\u003cp\u003eGenetic diversity index of \u003cem\u003eESR1\u003c/em\u003e gene in Cele Black sheep\u003c/p\u003e\n\u003cp\u003eThe differences of genotype frequency and allele frequency of different loci of \u003cem\u003eESR1\u003c/em\u003e gene in single lamb and multi-lamb of Cele Black sheep were shown in Table\u0026nbsp;\u003cspan class=\"InternalRef\"\u003e1\u003c/span\u003e, in which the genotype frequency and allele frequency of rs590747651 locus were statistically significant in single lamb and multi-lamb. As far as rs590747651 locus is concerned, AT is the dominant genotype, and the dominant allele in multi-lambs is A. Table\u0026nbsp;\u003cspan class=\"InternalRef\"\u003e2\u003c/span\u003e showed that the rs590747651 locus of \u003cem\u003eESR1\u003c/em\u003e gene was moderately polymorphic in both single lambs and multiple lambs of Black sheep (0.25\u0026thinsp;\u0026le;\u0026thinsp;PIC\u0026thinsp;\u0026lt;\u0026thinsp;0.5).\u003c/p\u003e\n\u003cp\u003eGenetic diversity index of \u003cem\u003eESR1\u003c/em\u003e gene in Hetian sheep\u003c/p\u003e\n\u003cp\u003eThe differences of genotype frequency and allele frequency of different loci of \u003cem\u003eESR1\u003c/em\u003e gene in single lamb and multi-lamb of Hetian sheep were shown in Table\u0026nbsp;\u003cspan class=\"InternalRef\"\u003e3\u003c/span\u003e, in which the genotype frequency and allele frequency of rs399356740 locus were statistically significant in single lamb and multi-lamb. In terms of rs399356740 loci, AT was the dominant genotype, T was the dominant allele in single lamb breeds, AA was the dominant genotype and A was the dominant allele in multi-lamb breeds. Table\u0026nbsp;\u003cspan class=\"InternalRef\"\u003e4\u003c/span\u003e showed that the rs399356740 locus of \u003cem\u003eESR1\u003c/em\u003e gene was moderately polymorphic in both single lambs and multiple lambs of Hetian sheep (0.25\u0026thinsp;\u0026le;\u0026thinsp;PIC\u0026thinsp;\u0026lt;\u0026thinsp;0.5).\u0026nbsp;\u003c/p\u003e\n\u003cdiv class=\"gridtable\"\u003e\n\u003ctable id=\"Tab1\" border=\"1\"\u003e\u003ccaption\u003e\n\u003cdiv class=\"CaptionNumber\"\u003eTable 1\u003c/div\u003e\n\u003cdiv class=\"CaptionContent\"\u003e\n\u003cp\u003eGene frequencies and genotype frequencies of different loci of \u003cem\u003eESR1\u003c/em\u003e gene in single and multiple lambs in Cele Black sheep.\u003c/p\u003e\n\u003c/div\u003e\n\u003c/caption\u003e\n\u003cthead\u003e\n\u003ctr\u003e\n\u003cth align=\"left\"\u003e\n\u003cp\u003eLocus\u003c/p\u003e\n\u003c/th\u003e\n\u003cth align=\"left\"\u003e\n\u003cp\u003eGenotype\u003c/p\u003e\n\u003c/th\u003e\n\u003cth align=\"left\"\u003e\n\u003cp\u003eGenotype frequency in uniparous sheep(sample size)\u003c/p\u003e\n\u003c/th\u003e\n\u003cth align=\"left\"\u003e\n\u003cp\u003eGenotype frequency in multiparous sheep(sample size)\u003c/p\u003e\n\u003c/th\u003e\n\u003cth align=\"left\"\u003e\n\u003cp\u003eP value\u003c/p\u003e\n\u003c/th\u003e\n\u003cth align=\"left\"\u003e\n\u003cp\u003eAllele\u003c/p\u003e\n\u003c/th\u003e\n\u003cth align=\"left\"\u003e\n\u003cp\u003eAllele frequency in uniparous sheep\u003c/p\u003e\n\u003c/th\u003e\n\u003cth align=\"left\"\u003e\n\u003cp\u003eAllele frequency in multiparous sheep\u003c/p\u003e\n\u003c/th\u003e\n\u003cth align=\"left\"\u003e\n\u003cp\u003eP value\u003c/p\u003e\n\u003c/th\u003e\n\u003c/tr\u003e\n\u003c/thead\u003e\n\u003ctbody\u003e\n\u003ctr\u003e\n\u003ctd rowspan=\"3\" align=\"left\"\u003e\n\u003cp\u003ers399356740\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"left\"\u003e\n\u003cp\u003eAA\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"char\" char=\".\"\u003e\n\u003cp\u003e0.27(13)\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"char\" char=\".\"\u003e\n\u003cp\u003e0.15(8)\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd rowspan=\"3\" align=\"char\" char=\".\"\u003e\n\u003cp\u003e0.06\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"left\"\u003e\n\u003cp\u003eA\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"char\" char=\".\"\u003e\n\u003cp\u003e0.43(21)\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"char\" char=\".\"\u003e\n\u003cp\u003e0.31(17)\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd rowspan=\"2\" align=\"char\" char=\".\"\u003e\n\u003cp\u003e0.23\u003c/p\u003e\n\u003c/td\u003e\n\u003c/tr\u003e\n\u003ctr\u003e\n\u003ctd align=\"left\"\u003e\n\u003cp\u003eAT\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"char\" char=\".\"\u003e\n\u003cp\u003e0.69(34)\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"char\" char=\".\"\u003e\n\u003cp\u003e0.85(46)\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"left\"\u003e\n\u003cp\u003eT\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"char\" char=\".\"\u003e\n\u003cp\u003e0.57(28)\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"char\" char=\".\"\u003e\n\u003cp\u003e0.69(37)\u003c/p\u003e\n\u003c/td\u003e\n\u003c/tr\u003e\n\u003ctr\u003e\n\u003ctd align=\"left\"\u003e\n\u003cp\u003eTT\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"char\" char=\".\"\u003e\n\u003cp\u003e0.04(2)\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"char\" char=\".\"\u003e\n\u003cp\u003e0.00(0)\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n\u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n\u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n\u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n\u003c/tr\u003e\n\u003ctr\u003e\n\u003ctd rowspan=\"3\" align=\"left\"\u003e\n\u003cp\u003ers590747651\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"left\"\u003e\n\u003cp\u003eAA\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"char\" char=\".\"\u003e\n\u003cp\u003e0.02(1)\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"char\" char=\".\"\u003e\n\u003cp\u003e0.00(0)\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd rowspan=\"3\" align=\"char\" char=\".\"\u003e\n\u003cp\u003e0.04\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"left\"\u003e\n\u003cp\u003eA\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"char\" char=\".\"\u003e\n\u003cp\u003e0.37(18)\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"char\" char=\".\"\u003e\n\u003cp\u003e0.59(32)\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd rowspan=\"2\" align=\"char\" char=\".\"\u003e\n\u003cp\u003e0.02\u003c/p\u003e\n\u003c/td\u003e\n\u003c/tr\u003e\n\u003ctr\u003e\n\u003ctd align=\"left\"\u003e\n\u003cp\u003eAT\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"char\" char=\".\"\u003e\n\u003cp\u003e0.92(45)\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"char\" char=\".\"\u003e\n\u003cp\u003e1.00(54)\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"left\"\u003e\n\u003cp\u003eT\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"char\" char=\".\"\u003e\n\u003cp\u003e0.63(31)\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"char\" char=\".\"\u003e\n\u003cp\u003e0.41(22)\u003c/p\u003e\n\u003c/td\u003e\n\u003c/tr\u003e\n\u003ctr\u003e\n\u003ctd align=\"left\"\u003e\n\u003cp\u003eTT\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"char\" char=\".\"\u003e\n\u003cp\u003e0.06(3)\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"char\" char=\".\"\u003e\n\u003cp\u003e0.00(0)\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n\u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n\u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n\u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n\u003c/tr\u003e\n\u003c/tbody\u003e\n\u003c/table\u003e\n\u003c/div\u003e\n\u003cp\u003eP\u0026thinsp;\u0026lt;\u0026thinsp;0.05 indicates the difference was significant.\u003c/p\u003e\n\u003cdiv class=\"gridtable\"\u003e\n\u003cdiv class=\"colspec\" align=\"left\"\u003e\u0026nbsp;\u003c/div\u003e\n\u003ctable id=\"Tab2\" border=\"1\"\u003e\u003ccaption\u003e\n\u003cdiv class=\"CaptionNumber\"\u003eTable 2\u003c/div\u003e\n\u003cdiv class=\"CaptionContent\"\u003e\n\u003cp\u003ePolymorphic information content, heterozygosity and effective alleles of different loci of \u003cem\u003eESR1\u003c/em\u003e gene in Cele Black sheep.\u003c/p\u003e\n\u003c/div\u003e\n\u003c/caption\u003e\n\u003cthead\u003e\n\u003ctr\u003e\n\u003cth align=\"left\"\u003e\n\u003cp\u003eLocus\u003c/p\u003e\n\u003c/th\u003e\n\u003cth align=\"left\"\u003e\n\u003cp\u003eHe in uniparous sheep\u003c/p\u003e\n\u003c/th\u003e\n\u003cth align=\"left\"\u003e\n\u003cp\u003eHe in multiparous sheep\u003c/p\u003e\n\u003c/th\u003e\n\u003cth align=\"left\"\u003e\n\u003cp\u003ePIC in uniparous sheep\u003c/p\u003e\n\u003c/th\u003e\n\u003cth align=\"left\"\u003e\n\u003cp\u003ePIC in multiparous sheep\u003c/p\u003e\n\u003c/th\u003e\n\u003cth align=\"left\"\u003e\n\u003cp\u003eNe in uniparous sheep\u003c/p\u003e\n\u003c/th\u003e\n\u003cth align=\"left\"\u003e\n\u003cp\u003eNe in multiparous sheep\u003c/p\u003e\n\u003c/th\u003e\n\u003c/tr\u003e\n\u003c/thead\u003e\n\u003ctbody\u003e\n\u003ctr\u003e\n\u003ctd align=\"left\"\u003e\n\u003cp\u003ers399356740\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"left\"\u003e\n\u003cp\u003e0.49\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"char\" char=\".\"\u003e\n\u003cp\u003e0.43\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"char\" char=\".\"\u003e\n\u003cp\u003e0.37\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"char\" char=\".\"\u003e\n\u003cp\u003e0.34\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"left\"\u003e\n\u003cp\u003e1.96\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"char\" char=\".\"\u003e\n\u003cp\u003e1.75\u003c/p\u003e\n\u003c/td\u003e\n\u003c/tr\u003e\n\u003ctr\u003e\n\u003ctd rowspan=\"3\" align=\"left\"\u003e\n\u003cp\u003ers590747651\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd rowspan=\"3\" align=\"left\"\u003e\n\u003cp\u003e0.47\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd rowspan=\"3\" align=\"char\" char=\".\"\u003e\n\u003cp\u003e0.48\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd rowspan=\"3\" align=\"char\" char=\".\"\u003e\n\u003cp\u003e0.36\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd rowspan=\"3\" align=\"char\" char=\".\"\u003e\n\u003cp\u003e0.37\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd rowspan=\"3\" align=\"left\"\u003e\n\u003cp\u003e1.87\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd rowspan=\"3\" align=\"char\" char=\".\"\u003e\n\u003cp\u003e1.94\u003c/p\u003e\n\u003c/td\u003e\n\u003c/tr\u003e\n\u003c/tbody\u003e\n\u003c/table\u003e\n\u003c/div\u003e\n\u003cdiv class=\"gridtable\"\u003e\n\u003cdiv class=\"colspec\" align=\"left\"\u003e\u0026nbsp;\u003c/div\u003e\n\u003cdiv class=\"colspec\" align=\"left\"\u003e\u0026nbsp;\u0026nbsp;\u003c/div\u003e\n\u003ctable id=\"Tab3\" border=\"1\"\u003e\u003ccaption\u003e\n\u003cdiv class=\"CaptionNumber\"\u003eTable 3\u003c/div\u003e\n\u003cdiv class=\"CaptionContent\"\u003e\n\u003cp\u003eGene frequencies and genotype frequencies of different loci of \u003cem\u003eESR1\u003c/em\u003e gene in single and multiple lambs in Hetian sheep.\u003c/p\u003e\n\u003c/div\u003e\n\u003c/caption\u003e\n\u003cthead\u003e\n\u003ctr\u003e\n\u003cth align=\"left\"\u003e\n\u003cp\u003eLocus\u003c/p\u003e\n\u003c/th\u003e\n\u003cth align=\"left\"\u003e\n\u003cp\u003eGenotype\u003c/p\u003e\n\u003c/th\u003e\n\u003cth align=\"left\"\u003e\n\u003cp\u003eGenotype frequency in uniparous sheep(sample size)\u003c/p\u003e\n\u003c/th\u003e\n\u003cth align=\"left\"\u003e\n\u003cp\u003eGenotype frequency in multiparous sheep(sample size)\u003c/p\u003e\n\u003c/th\u003e\n\u003cth align=\"left\"\u003e\n\u003cp\u003eP value\u003c/p\u003e\n\u003c/th\u003e\n\u003cth align=\"left\"\u003e\n\u003cp\u003eAllele\u003c/p\u003e\n\u003c/th\u003e\n\u003cth align=\"left\"\u003e\n\u003cp\u003eAllele frequency in uniparous sheep\u003c/p\u003e\n\u003c/th\u003e\n\u003cth align=\"left\"\u003e\n\u003cp\u003eAllele frequency in multiparous sheep\u003c/p\u003e\n\u003c/th\u003e\n\u003cth align=\"left\"\u003e\n\u003cp\u003eP value\u003c/p\u003e\n\u003c/th\u003e\n\u003c/tr\u003e\n\u003c/thead\u003e\n\u003ctbody\u003e\n\u003ctr\u003e\n\u003ctd rowspan=\"3\" align=\"left\"\u003e\n\u003cp\u003ers399356740\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"left\"\u003e\n\u003cp\u003eAA\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"char\" char=\".\"\u003e\n\u003cp\u003e0.34(21)\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"char\" char=\".\"\u003e\n\u003cp\u003e0.56(24)\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd rowspan=\"3\" align=\"char\" char=\".\"\u003e\n\u003cp\u003e0.04\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"left\"\u003e\n\u003cp\u003eA\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"char\" char=\".\"\u003e\n\u003cp\u003e0.37(23)\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"char\" char=\".\"\u003e\n\u003cp\u003e0.60(26)\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd rowspan=\"2\" align=\"char\" char=\".\"\u003e\n\u003cp\u003e0.02\u003c/p\u003e\n\u003c/td\u003e\n\u003c/tr\u003e\n\u003ctr\u003e\n\u003ctd align=\"left\"\u003e\n\u003cp\u003eAT\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"char\" char=\".\"\u003e\n\u003cp\u003e0.61(38)\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"char\" char=\".\"\u003e\n\u003cp\u003e0.44(19)\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"left\"\u003e\n\u003cp\u003eT\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"char\" char=\".\"\u003e\n\u003cp\u003e0.63(39)\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"char\" char=\".\"\u003e\n\u003cp\u003e0.40(17)\u003c/p\u003e\n\u003c/td\u003e\n\u003c/tr\u003e\n\u003ctr\u003e\n\u003ctd align=\"left\"\u003e\n\u003cp\u003eTT\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"char\" char=\".\"\u003e\n\u003cp\u003e0.05(3)\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"char\" char=\".\"\u003e\n\u003cp\u003e0.00(0)\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n\u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n\u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n\u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n\u003c/tr\u003e\n\u003ctr\u003e\n\u003ctd rowspan=\"3\" align=\"left\"\u003e\n\u003cp\u003ers590747651\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"left\"\u003e\n\u003cp\u003eAA\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"char\" char=\".\"\u003e\n\u003cp\u003e0.21(13)\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"char\" char=\".\"\u003e\n\u003cp\u003e0.39(17)\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd rowspan=\"3\" align=\"char\" char=\".\"\u003e\n\u003cp\u003e0.05\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"left\"\u003e\n\u003cp\u003eA\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"char\" char=\".\"\u003e\n\u003cp\u003e0.66(41)\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"char\" char=\".\"\u003e\n\u003cp\u003e0.65(28)\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd rowspan=\"2\" align=\"char\" char=\".\"\u003e\n\u003cp\u003e0.27\u003c/p\u003e\n\u003c/td\u003e\n\u003c/tr\u003e\n\u003ctr\u003e\n\u003ctd align=\"left\"\u003e\n\u003cp\u003eAT\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"char\" char=\".\"\u003e\n\u003cp\u003e0.50(31)\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"char\" char=\".\"\u003e\n\u003cp\u003e0.28(12)\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"left\"\u003e\n\u003cp\u003eT\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"char\" char=\".\"\u003e\n\u003cp\u003e0.34(21)\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"char\" char=\".\"\u003e\n\u003cp\u003e0.35(15)\u003c/p\u003e\n\u003c/td\u003e\n\u003c/tr\u003e\n\u003ctr\u003e\n\u003ctd align=\"left\"\u003e\n\u003cp\u003eTT\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"char\" char=\".\"\u003e\n\u003cp\u003e0.29(18)\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"char\" char=\".\"\u003e\n\u003cp\u003e0.33(14)\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n\u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n\u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n\u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n\u003c/tr\u003e\n\u003c/tbody\u003e\n\u003c/table\u003e\n\u003c/div\u003e\n\u003cp\u003eP\u0026thinsp;\u0026lt;\u0026thinsp;0.05 indicates the difference was significant.\u0026nbsp;\u003c/p\u003e\n\u003cdiv class=\"gridtable\"\u003e\n\u003ctable id=\"Tab4\" border=\"1\"\u003e\u003ccaption\u003e\n\u003cdiv class=\"CaptionNumber\"\u003eTable 4\u003c/div\u003e\n\u003cdiv class=\"CaptionContent\"\u003e\n\u003cp\u003ePolymorphic information content, heterozygosity and effective alleles of \u003cem\u003eESR1\u003c/em\u003e gene at different loci of \u003cem\u003eESR1\u003c/em\u003e gene in Hetian sheep.\u003c/p\u003e\n\u003c/div\u003e\n\u003c/caption\u003e\n\u003cthead\u003e\n\u003ctr\u003e\n\u003cth align=\"left\"\u003e\n\u003cp\u003eLocus\u003c/p\u003e\n\u003c/th\u003e\n\u003cth align=\"left\"\u003e\n\u003cp\u003eHe in uniparous sheep\u003c/p\u003e\n\u003c/th\u003e\n\u003cth align=\"left\"\u003e\n\u003cp\u003eHe in multiparous sheep\u003c/p\u003e\n\u003c/th\u003e\n\u003cth align=\"left\"\u003e\n\u003cp\u003ePIC in uniparous sheep\u003c/p\u003e\n\u003c/th\u003e\n\u003cth align=\"left\"\u003e\n\u003cp\u003ePIC in multiparous sheep\u003c/p\u003e\n\u003c/th\u003e\n\u003cth align=\"left\"\u003e\n\u003cp\u003eNe in uniparous sheep\u003c/p\u003e\n\u003c/th\u003e\n\u003cth align=\"left\"\u003e\n\u003cp\u003eNe in multiparous sheep\u003c/p\u003e\n\u003c/th\u003e\n\u003c/tr\u003e\n\u003c/thead\u003e\n\u003ctbody\u003e\n\u003ctr\u003e\n\u003ctd align=\"left\"\u003e\n\u003cp\u003ers399356740\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"char\" char=\".\"\u003e\n\u003cp\u003e0.47\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"char\" char=\".\"\u003e\n\u003cp\u003e0.48\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"char\" char=\".\"\u003e\n\u003cp\u003e0.36\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"char\" char=\".\"\u003e\n\u003cp\u003e0.36\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"char\" char=\".\"\u003e\n\u003cp\u003e1.87\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"char\" char=\".\"\u003e\n\u003cp\u003e1.92\u003c/p\u003e\n\u003c/td\u003e\n\u003c/tr\u003e\n\u003ctr\u003e\n\u003ctd align=\"left\"\u003e\n\u003cp\u003ers590747651\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"char\" char=\".\"\u003e\n\u003cp\u003e0.45\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"char\" char=\".\"\u003e\n\u003cp\u003e0.46\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"char\" char=\".\"\u003e\n\u003cp\u003e0.35\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"char\" char=\".\"\u003e\n\u003cp\u003e0.35\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"char\" char=\".\"\u003e\n\u003cp\u003e1.81\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"char\" char=\".\"\u003e\n\u003cp\u003e1.83\u003c/p\u003e\n\u003c/td\u003e\n\u003c/tr\u003e\n\u003c/tbody\u003e\n\u003c/table\u003e\n\u003c/div\u003e\n\u003cp\u003ePopulation genetic analysis of different SNPs of \u003cem\u003eESR1\u003c/em\u003e gene in different sheep breeds\u003c/p\u003e\n\u003cp\u003eThe loci of rs399356740 and rs590747651 of \u003cem\u003eESR1\u003c/em\u003e gene were moderately polymorphic in Cele Black sheep and Hetian sheep (0.25\u0026thinsp;\u0026lt;\u0026thinsp;PIC\u0026thinsp;\u0026lt;\u0026thinsp;0.5). Chi-square test showed that the \u003cem\u003eESR1\u003c/em\u003e gene rs399356740 locus was in Hardy-Weinberg disequilibrium in Cele Black sheep and Hetian sheep breeds, and the rs590747651 locus in \u003cem\u003eESR1\u003c/em\u003e gene was in Hardy-Weinberg disequilibrium in Cele Black sheep and was in Hardy-Weinberg equilibrium in Hetian sheep. Cele Black sheep is a high yield sheep, but Hetian sheep is not. in the two loci of \u003cem\u003eESR1\u003c/em\u003e gene rs399356740 and rs590747651, the dominant allele of rs399356740 in the two sheep breeds is T (Table\u0026nbsp;\u003cspan class=\"InternalRef\"\u003e5\u003c/span\u003e), but the dominant allele of rs590747651 in Cele Black sheep is T, and the dominant allele in Hetian sheep is A, so it is speculated that this may be the reason for the difference in fecundity between the two sheep populations.\u003c/p\u003e\n\u003cdiv class=\"gridtable\"\u003e\n\u003ctable id=\"Tab5\" border=\"1\"\u003e\u003ccaption\u003e\n\u003cdiv class=\"CaptionNumber\"\u003eTable 5\u003c/div\u003e\n\u003cdiv class=\"CaptionContent\"\u003e\n\u003cp\u003ePopulation genetic analysis of different loci of \u003cem\u003eESR1\u003c/em\u003e gene in different sheep breeds.\u003c/p\u003e\n\u003c/div\u003e\n\u003c/caption\u003e\n\u003cthead\u003e\n\u003ctr\u003e\n\u003cth align=\"left\"\u003e\n\u003cp\u003eLocus\u003c/p\u003e\n\u003c/th\u003e\n\u003cth align=\"left\"\u003e\n\u003cp\u003eVariety\u003c/p\u003e\n\u003c/th\u003e\n\u003cth colspan=\"3\" align=\"left\"\u003e\n\u003cp\u003eGenotype frequency(sample size)\u003c/p\u003e\n\u003c/th\u003e\n\u003cth colspan=\"2\" align=\"left\"\u003e\n\u003cp\u003eAllele\u003c/p\u003e\n\u003cp\u003eFrequency\u003c/p\u003e\n\u003c/th\u003e\n\u003cth align=\"left\"\u003e\n\u003cp\u003ePolymorphism information content(PIC)\u003c/p\u003e\n\u003c/th\u003e\n\u003cth align=\"left\"\u003e\n\u003cp\u003eHeterozy gosity\u003c/p\u003e\n\u003cp\u003e(He)\u003c/p\u003e\n\u003c/th\u003e\n\u003cth align=\"left\"\u003e\n\u003cp\u003eEffective number of allele(Ne)\u003c/p\u003e\n\u003c/th\u003e\n\u003cth align=\"left\"\u003e\n\u003cp\u003eP value\u003c/p\u003e\n\u003c/th\u003e\n\u003c/tr\u003e\n\u003c/thead\u003e\n\u003ctbody\u003e\n\u003ctr\u003e\n\u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n\u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n\u003ctd align=\"left\"\u003e\n\u003cp\u003eAA\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"left\"\u003e\n\u003cp\u003eAT\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"left\"\u003e\n\u003cp\u003eTT\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"left\"\u003e\n\u003cp\u003eA\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"left\"\u003e\n\u003cp\u003eT\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n\u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n\u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n\u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n\u003c/tr\u003e\n\u003ctr\u003e\n\u003ctd rowspan=\"2\" align=\"left\"\u003e\n\u003cp\u003ers399356740\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"left\"\u003e\n\u003cp\u003eCele Black sheep\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"left\"\u003e\n\u003cp\u003e0.20(21)\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"left\"\u003e\n\u003cp\u003e0.78(80)\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"left\"\u003e\n\u003cp\u003e0.02(2)\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"left\"\u003e\n\u003cp\u003e0.37\u003c/p\u003e\n\u003cp\u003e(38)\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"left\"\u003e\n\u003cp\u003e0.63\u003c/p\u003e\n\u003cp\u003e(65)\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"char\" char=\".\"\u003e\n\u003cp\u003e0.36\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"char\" char=\".\"\u003e\n\u003cp\u003e0.47\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"char\" char=\".\"\u003e\n\u003cp\u003e1.87\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"char\" char=\".\"\u003e\n\u003cp\u003e0.00\u003c/p\u003e\n\u003c/td\u003e\n\u003c/tr\u003e\n\u003ctr\u003e\n\u003ctd align=\"left\"\u003e\n\u003cp\u003eHetian sheep\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"left\"\u003e\n\u003cp\u003e0.43(45)\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"left\"\u003e\n\u003cp\u003e0.54(57)\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"left\"\u003e\n\u003cp\u003e0.03(3)\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"left\"\u003e\n\u003cp\u003e0.47\u003c/p\u003e\n\u003cp\u003e(49)\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"left\"\u003e\n\u003cp\u003e0.53\u003c/p\u003e\n\u003cp\u003e(56)\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"char\" char=\".\"\u003e\n\u003cp\u003e0.37\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"char\" char=\".\"\u003e\n\u003cp\u003e0.50\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"char\" char=\".\"\u003e\n\u003cp\u003e1.99\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"char\" char=\".\"\u003e\n\u003cp\u003e0.00\u003c/p\u003e\n\u003c/td\u003e\n\u003c/tr\u003e\n\u003ctr\u003e\n\u003ctd rowspan=\"2\" align=\"left\"\u003e\n\u003cp\u003ers590747651\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"left\"\u003e\n\u003cp\u003eCele Black sheep\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"left\"\u003e\n\u003cp\u003e0.01(1)\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"left\"\u003e\n\u003cp\u003e0.96(99)\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"left\"\u003e\n\u003cp\u003e0.03(3)\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"left\"\u003e\n\u003cp\u003e0.49\u003c/p\u003e\n\u003cp\u003e(50)\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"left\"\u003e\n\u003cp\u003e0.51\u003c/p\u003e\n\u003cp\u003e(53)\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"char\" char=\".\"\u003e\n\u003cp\u003e0.37\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"char\" char=\".\"\u003e\n\u003cp\u003e0.50\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"char\" char=\".\"\u003e\n\u003cp\u003e1.99\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"char\" char=\".\"\u003e\n\u003cp\u003e0.00\u003c/p\u003e\n\u003c/td\u003e\n\u003c/tr\u003e\n\u003ctr\u003e\n\u003ctd align=\"left\"\u003e\n\u003cp\u003eHetian sheep\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"left\"\u003e\n\u003cp\u003e0.29(30)\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"left\"\u003e\n\u003cp\u003e0.41(43)\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"left\"\u003e\n\u003cp\u003e0.30(32)\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"left\"\u003e\n\u003cp\u003e0.66\u003c/p\u003e\n\u003cp\u003e(69)\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"left\"\u003e\n\u003cp\u003e0.34\u003c/p\u003e\n\u003cp\u003e(36)\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"char\" char=\".\"\u003e\n\u003cp\u003e0.35\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"char\" char=\".\"\u003e\n\u003cp\u003e0.45\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"char\" char=\".\"\u003e\n\u003cp\u003e1.81\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"char\" char=\".\"\u003e\n\u003cp\u003e0.06\u003c/p\u003e\n\u003c/td\u003e\n\u003c/tr\u003e\n\u003c/tbody\u003e\n\u003c/table\u003e\n\u003c/div\u003e\n\u003cp\u003eP\u0026thinsp;\u0026lt;\u0026thinsp;0.05 indicates the difference was significant.\u003c/p\u003e\n\u003cp\u003eAnalysis of the correlation between SNP locus of \u003cem\u003eESR1\u003c/em\u003e gene and litter size\u003c/p\u003e\n\u003cdiv class=\"gridtable\"\u003e\n\u003cdiv class=\"colspec\" align=\"left\"\u003e\u0026nbsp;\n\u003cp\u003eTable 6 showed that there was no significant correlation between rs399356740 genotypes of \u003cem\u003eESR1\u003c/em\u003e gene and litter size in Cele Black sheep and Hetian sheep, and there was no significant correlation between genotypes of \u003cem\u003eESR1\u003c/em\u003e gene rs590747651 locus and litter size of Cele Black sheep. The lambing number of AA and TT genotypes of \u003cem\u003eESR1\u003c/em\u003e gene rs590747651 mutation in Hetian sheep was significantly higher than that of AT type, and there was no significant difference between AA and TT genotypes.\u003c/p\u003e\n\u003c/div\u003e\n\u003cdiv class=\"colspec\" align=\"left\"\u003e\u0026nbsp;\u003c/div\u003e\n\u003cdiv class=\"colspec\" align=\"left\"\u003e\u0026nbsp;\u003c/div\u003e\n\u003cdiv class=\"colspec\" align=\"left\"\u003e\u0026nbsp;\u003c/div\u003e\n\u003ctable id=\"Tab6\" border=\"1\"\u003e\u003ccaption\u003e\n\u003cdiv class=\"CaptionNumber\"\u003eTable 6\u003c/div\u003e\n\u003cdiv class=\"CaptionContent\"\u003e\n\u003cp\u003eCorrelation Analysis of SNP Locus of \u003cem\u003eESR1\u003c/em\u003e Gene and litter size of Cele Black sheep and Hetian sheep.\u003c/p\u003e\n\u003c/div\u003e\n\u003c/caption\u003e\n\u003cthead\u003e\n\u003ctr\u003e\n\u003cth align=\"left\"\u003e\n\u003cp\u003eVariety\u003c/p\u003e\n\u003c/th\u003e\n\u003cth align=\"left\"\u003e\n\u003cp\u003eSNP\u003c/p\u003e\n\u003c/th\u003e\n\u003cth align=\"left\"\u003e\n\u003cp\u003eGenotype\u003c/p\u003e\n\u003c/th\u003e\n\u003cth align=\"left\"\u003e\n\u003cp\u003eNumber of lambs\u003c/p\u003e\n\u003c/th\u003e\n\u003c/tr\u003e\n\u003c/thead\u003e\n\u003ctbody\u003e\n\u003ctr\u003e\n\u003ctd rowspan=\"6\" align=\"left\"\u003e\n\u003cp\u003eCele Black sheep\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd rowspan=\"3\" align=\"left\"\u003e\n\u003cp\u003ers399356740\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"left\"\u003e\n\u003cp\u003eAA\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"left\"\u003e\n\u003cp\u003e1.48\u0026thinsp;\u0026plusmn;\u0026thinsp;0.50\u003c/p\u003e\n\u003c/td\u003e\n\u003c/tr\u003e\n\u003ctr\u003e\n\u003ctd align=\"left\"\u003e\n\u003cp\u003eAT\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"left\"\u003e\n\u003cp\u003e1.50\u0026thinsp;\u0026plusmn;\u0026thinsp;0.50\u003c/p\u003e\n\u003c/td\u003e\n\u003c/tr\u003e\n\u003ctr\u003e\n\u003ctd align=\"left\"\u003e\n\u003cp\u003eTT\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"left\"\u003e\n\u003cp\u003e1.00\u0026thinsp;\u0026plusmn;\u0026thinsp;0.00\u003c/p\u003e\n\u003c/td\u003e\n\u003c/tr\u003e\n\u003ctr\u003e\n\u003ctd rowspan=\"3\" align=\"left\"\u003e\n\u003cp\u003ers590747651\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"left\"\u003e\n\u003cp\u003eAA\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"left\"\u003e\n\u003cp\u003e1.00\u0026thinsp;\u0026plusmn;\u0026thinsp;0.00\u003c/p\u003e\n\u003c/td\u003e\n\u003c/tr\u003e\n\u003ctr\u003e\n\u003ctd align=\"left\"\u003e\n\u003cp\u003eAT\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"left\"\u003e\n\u003cp\u003e1.55\u0026thinsp;\u0026plusmn;\u0026thinsp;0.50\u003c/p\u003e\n\u003c/td\u003e\n\u003c/tr\u003e\n\u003ctr\u003e\n\u003ctd align=\"left\"\u003e\n\u003cp\u003eTT\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"left\"\u003e\n\u003cp\u003e1.00\u0026thinsp;\u0026plusmn;\u0026thinsp;0.00\u003c/p\u003e\n\u003c/td\u003e\n\u003c/tr\u003e\n\u003ctr\u003e\n\u003ctd rowspan=\"5\" align=\"left\"\u003e\n\u003cp\u003eHetian sheep\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd rowspan=\"3\" align=\"left\"\u003e\n\u003cp\u003ers399356740\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"left\"\u003e\n\u003cp\u003eAA\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"left\"\u003e\n\u003cp\u003e1.53\u0026thinsp;\u0026plusmn;\u0026thinsp;0.50\u003c/p\u003e\n\u003c/td\u003e\n\u003c/tr\u003e\n\u003ctr\u003e\n\u003ctd align=\"left\"\u003e\n\u003cp\u003eAT\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"left\"\u003e\n\u003cp\u003e1.33\u0026thinsp;\u0026plusmn;\u0026thinsp;0.48\u003c/p\u003e\n\u003c/td\u003e\n\u003c/tr\u003e\n\u003ctr\u003e\n\u003ctd align=\"left\"\u003e\n\u003cp\u003eTT\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"left\"\u003e\n\u003cp\u003e1.00\u0026thinsp;\u0026plusmn;\u0026thinsp;0.00\u003c/p\u003e\n\u003c/td\u003e\n\u003c/tr\u003e\n\u003ctr\u003e\n\u003ctd rowspan=\"2\" align=\"left\"\u003e\n\u003cp\u003ers590747651\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"left\"\u003e\n\u003cp\u003eAA\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"left\"\u003e\n\u003cp\u003e1.57\u0026thinsp;\u0026plusmn;\u0026thinsp;0.50 \u003csup\u003ea\u003c/sup\u003e\u003c/p\u003e\n\u003c/td\u003e\n\u003c/tr\u003e\n\u003ctr\u003e\n\u003ctd align=\"left\"\u003e\n\u003cp\u003eAT\u003c/p\u003e\n\u003c/td\u003e\n\u003ctd align=\"left\"\u003e\n\u003cp\u003e1.28\u0026thinsp;\u0026plusmn;\u0026thinsp;0.45 \u003csup\u003eb\u003c/sup\u003e\u003c/p\u003e\n\u003c/td\u003e\n\u003c/tr\u003e\n\u003c/tbody\u003e\n\u003c/table\u003e\n\u003c/div\u003e\n\u003cdiv class=\"BlockQuote\"\u003e\n\u003cp\u003e\u003cstrong\u003eNote:\u0026nbsp;\u003c/strong\u003ewhen comparing different genotypes, the difference is significant when there is any letter difference after the data (P\u0026thinsp;\u0026lt;\u0026thinsp;0.05).\u003c/p\u003e\n\u003c/div\u003e"},{"header":"4 Discussion","content":"\u003cp\u003eCele Black sheep and Hetian sheep are two local sheep breeds unique to Xinjiang, China, and their reproductive performance is quite different, among which the lambing rate of Hetian sheep is low, which seriously hinders the economic development of the local sheep industry. However, Hetian sheep, as a characteristic sheep germplasm resource in Xinjiang, has formed the germplasm characteristics of drought and heat tolerance and strong adaptability under the unique natural ecological conditions of Hetian region, Ewes can be in estrus all the year round, and the lambing rate of Cele Black sheep is high. Therefore, the breeding of Cele Black sheep and Hetian sheep plays an important role in promoting the overall sheep breeding and economic development in Xinjiang [10]. Bai Shu et al (2013) found that \u003cem\u003eESR1\u003c/em\u003e was expressed in uterine lumen epithelium, glandular epithelium, matrix, vascular endothelium and smooth muscle cell nucleus in Jining grey goats, which confirmed that \u003cem\u003eESR1\u003c/em\u003e was involved in regulating the proliferation and differentiation of endometrium and myometrium [11].After knocking out the ESR gene of female mice, ESR-deficient female mice appeared luteinizing hormone regulation disorder and ovary anovulation, which affected the fertility of mice [7]. Overexpression of \u003cem\u003eESR1\u003c/em\u003e may lead to follicular atresia [12]. Generally speaking, \u003cem\u003eESR1\u003c/em\u003e may have a certain effect on reproductive performance, and \u003cem\u003eESR1\u003c/em\u003e gene, as one of the subtypes of ESR, plays an important role in lambing performance of mammals. In early experiments, it has been confirmed that \u003cem\u003eESR1\u003c/em\u003e in endometrium is necessary to identify pregnancy and maternal development in pigs [13, 14]. Rempel et al found that the SNP loci of \u003cem\u003eESR1\u003c/em\u003e and \u003cem\u003eESR2\u003c/em\u003e were significantly correlated with litter size. The high yield genotype of ESR gene Pvu II restriction site of Baoshan pig was AB type, and the B allele had a positive effect on the litter size of Baoshan sows. Shun Wu et al found that the TT genotype of ESR g.C1665T locus and the AA genotype of g.A1755G locus could significantly increase the total litter size of the first birth of American large White pigs [15\u0026ndash;17]. There are many studies of \u003cem\u003eESR1\u003c/em\u003e gene on the reproductive performance of other mammals, such as duck [18], quail [19] and fish [20]. These studies show that \u003cem\u003eESR1\u003c/em\u003e gene plays a key role in estrus ovulation and lambing. The purpose of this study is to identify \u003cem\u003eESR1\u003c/em\u003e gene as a potential molecular marker and further study the relationship between MAS and lambing, so as to apply MAS to sheep breeding.\u003c/p\u003e\u003cp\u003eIt was reported that there was a significant correlation between the \u003cem\u003eESR1\u003c/em\u003e locus rs399356740 locus and the number of Small-Tailed Han lambs, and the number of lambs in AA ewes was significantly higher than that in TT ewes, but there was no significant correlation between rs590747651 locus and the number of lambs in different small-tailed Han sheep, that is, there was more and more evidence that mutations in \u003cem\u003eESR1\u003c/em\u003e gene locus were related to animal reproduction [21]. The current study aimed to check the association of \u003cem\u003eESR1\u003c/em\u003e rs399356740 and rs590747651 with the litter size in the Cele Black sheep and Hetian sheep. We found that the rs399356740 and rs590747651 loci of \u003cem\u003eESR1\u003c/em\u003e gene in Cele Black sheep and Hetian sheep were moderately polymorphic. Except that the rs590747651 locus was in Hardy-Weinberg equilibrium in Hetian sheep, none of them showed Hardy-Weinberg equilibrium, which may be due to site imbalance caused by natural selection or artificial intervention, or due to the small number of sheep involved in this study. Our study shows that the rs399356740 and Cele Black sheep of \u003cem\u003eESR1\u003c/em\u003e gene have nothing to do with the litter size of Hetian sheep, and rs590747651 has nothing to do with the litter size of Cele Black sheep, which may be caused by the lack of high selection in the breeding process, but it can not be ruled out that it is due to the small sample size. However, there was a significant correlation between rs590747651 and lambing number of Hetian sheep. The lambing number of AA and TT type ewes was significantly higher than that of AT type ewes. Therefore, in the process of Hetian sheep breeding, homozygous individuals of AA and TT can be retained to improve the fecundity of Hetian sheep. This study provided a preliminary reference for marker-assisted selection of litter size in Hetian sheep, but in view of the shortage of samples, the TT genotypes of the detected rs399356740 loci in Cele Black sheep and Hetian sheep, and the number of AA and TT genotypes of rs590747651 in Cele Black sheep were insufficient, it is necessary to expand the sample size and improve the accuracy of the experiment.\u003c/p\u003e"},{"header":"5 Conclusions","content":"\u003cp\u003eThis study summarized the association between \u003cem\u003eESR1\u003c/em\u003e gene rs590747651 and rs399356740 and lambing number of Cele black sheep and Hetian sheep, and found that the variation of rs399356740 was related to the litter size of Hetian sheep. Statistical analysis showed that the litter size of \u003cem\u003eESR1\u003c/em\u003e rs399356740 homozygous genotype was significantly higher than that of heterozygous genotype.\u003c/p\u003e"},{"header":"Abbreviations","content":"\u003ctable border=\"1\" cellspacing=\"0\" cellpadding=\"0\"\u003e\n \u003ctbody\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 50%;\"\u003e\n \u003cp\u003e\u003cstrong\u003eItem\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 50%;\"\u003e\n \u003cp\u003e\u003cstrong\u003eUnit\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 50%;\"\u003e\n \u003cp\u003ePCR\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 50%;\"\u003e\n \u003cp\u003ePolymerase Chain Reaction\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 50%;\"\u003e\n \u003cp\u003eESR\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 50%;\"\u003e\n \u003cp\u003eEstrogen Receptor\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 50%;\"\u003e\n \u003cp\u003eGDF9\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 50%;\"\u003e\n \u003cp\u003eGrowth Differentiation Factor 9\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 50%;\"\u003e\n \u003cp\u003eBMP15\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 50%;\"\u003e\n \u003cp\u003eBone Morphogenetic Protein 15\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 50%;\"\u003e\n \u003cp\u003eBMPR1B\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 50%;\"\u003e\n \u003cp\u003eBone Morphogenetic Protein Receptor 1B\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 50%;\"\u003e\n \u003cp\u003eESR1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 50%;\"\u003e\n \u003cp\u003eEstrogen Receptor 1\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 50%;\"\u003e\n \u003cp\u003eGnRH\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 50%;\"\u003e\n \u003cp\u003eGonadotropin-Releasing Hormone\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 50%;\"\u003e\n \u003cp\u003eLH\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 50%;\"\u003e\n \u003cp\u003eLuteinizing Hormone\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 50%;\"\u003e\n \u003cp\u003eSNP\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 50%;\"\u003e\n \u003cp\u003eSingle Nucleotide Polymorphism\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003c/tbody\u003e\n\u003c/table\u003e"},{"header":"Declarations","content":"\u003ch2\u003e7.1\u0026nbsp; \u0026nbsp; \u0026nbsp;Ethics approval and consent to participate\u003c/h2\u003e\n\u003cp\u003eFor experimental animals, all protocols were performed in accordance with the\u0026nbsp;\u0026ldquo;Guide to Animal Experimentation\u0026rdquo;\u0026nbsp;and approved by the Use Committee under the norms of the Ethics Committee of Tarim University of Science and Technology (SYXK 2020-009).\u003c/p\u003e\n\u003ch2\u003e7.2\u0026nbsp; \u0026nbsp; \u0026nbsp;Consent for publication\u003c/h2\u003e\n\u003cp\u003eNot applicable.\u003c/p\u003e\n\u003cp\u003eThis manuscript does not contain personal data from any individual. All authors have reviewed the final version of the manuscript and consent to its publication in BMC Genomics.\u003c/p\u003e\n\u003ch2\u003e7.3\u0026nbsp; \u0026nbsp; \u0026nbsp;Availability of data and materials\u003c/h2\u003e\n\u003cp\u003eThe datasets generated and analyzed during this study are included in this published article and its supplementary files. The raw sequencing data are available from the corresponding author upon reasonable request.\u003c/p\u003e\n\u003cp\u003eBiological materials: Blood samples from Cele Black sheep and Hetian sheep are preserved at the Key Laboratory of Utilization of Surrounding Tarim Livestock and Grass Resources (Tarim University) and may be requested for non-commercial research purposes.\u003c/p\u003e\n\u003cp\u003ePublic data resources:\u0026nbsp;Reference sequences for the \u003cem\u003eESR1\u003c/em\u003e gene (NCBI Accession: NC_056061.1) and SNP loci (rs399356740, rs590747651) were obtained from the NCBI (https://www.ncbi.nlm.nih.gov/) and Ensembl (https://www.ensembl.org/) databases.\u003c/p\u003e\n\u003cp\u003ePrimers: All primer sequences used for PCR amplification are provided in the Methods section.\u003c/p\u003e\n\u003ch2\u003e7.4\u0026nbsp; \u0026nbsp; \u0026nbsp;Competing interests.\u0026nbsp;\u003c/h2\u003e\n\u003cp\u003eThe authors declare that this research was conducted in the absence of any conflicts of interest.\u003c/p\u003e\n\u003ch2\u003e7.5\u0026nbsp; \u0026nbsp; \u0026nbsp;Funding.\u003c/h2\u003e\n\u003cp\u003e\u0026nbsp;The study was made possible thanks to funding from the Xinjiang Production and Construction Corps (Grant/Award Numbers: 2022CB00104) and the President\u0026apos;s Fund Project of Tarim University (Grant/Award Numbers: TDZKCX202401).\u003c/p\u003e\n\u003ch2\u003e7.6\u0026nbsp; \u0026nbsp; \u0026nbsp;Author Contributions.\u0026nbsp;\u003c/h2\u003e\n\u003cp\u003eQ.H., G.R conceived the study, conducted data analysis, and prepared the numbers and tables. Q.H., G.R. and W.L. collected the samples, L.Z, and H.S. conducted genetic diversity analyses F.X. supervised the study. Q.H. This manuscript has been prepared. All the authors have read and agreed to the published version of the manuscript.\u003c/p\u003e"},{"header":"References","content":"\u003col\u003e\u003cli\u003e\u003cspan\u003e Yue C, Bai WL, Zheng YY, Hui TY, Sun JM, Guo D, Guo SL. Correlation. Analysis of candidate gene SNP for high-yield in Liaoning cashmere goats with litter size and cashmere performance. Anim Biotechnol 2021;32:43\u0026ndash;50. https://doi.org/10.1080/10495398\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003e Shi HC, Bai J, Niu ZG, Muniresha, Fen LJ, Jia B. Study on Candidate Gene for Fecundity Traits in Xingjiang Cele Black Sheep. Afr J Biotechnol 2010;9:8498\u0026ndash;850. https://doi.org/10.5897/AJB10.1003\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003e Hasimu A, Maitusong A. Survey and analysis of the development trend of Hetian sheep in Yutian County. Xinjiang xumuye 2013;S2:20\u0026ndash;21. https://doi.org/10.16795/j.cnki.xjxmy\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003e Miao X, Luo Q. Genome-wide transcriptome analysis between small-tail Han sheep and the Surabaya fur sheep using high-throughput RNA sequencing. Reprod 2013;145:587\u0026thinsp;\u0026minus;\u0026thinsp;96. https://doi.org/10.1530/REP-12-0507\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003e Dorling AA, Todman MG, Korach KS, Herbison AE. Critical role for estrogen receptor alpha in negative feedback regulation of gonadotropin-releasing hormone mRNA expression in the female mouse. 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Chin J Anim Sci 2018;54:36\u0026ndash;40. https://doi.org/10.19556/j.0258-7033.2018-10-036\u003c/span\u003e\u003c/li\u003e\u003c/ol\u003e"}],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":true,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":false,"hideJournal":true,"highlight":"","institution":"","isAcceptedByJournal":false,"isAuthorSuppliedPdf":false,"isDeskRejected":"","isHiddenFromSearch":false,"isInQc":false,"isInWorkflow":false,"isPdf":false,"isPdfUpToDate":true,"isWithdrawnOrRetracted":false,"journal":{"display":true,"email":"[email protected]","identity":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true},"keywords":"ESR1 gene, polymorphism, lamb, Cele Black sheep, Hetian sheep","lastPublishedDoi":"10.21203/rs.3.rs-7326024/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-7326024/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"\u003cp\u003eLitter size is one of the most important traits of sheep. The purpose of this study is to detect the \u003cem\u003eESR1\u003c/em\u003e gene in Cele Black sheep and Hetian sheep populations, so as to lay a foundation for further improving the litter size of the population and establishing a core population with high fecundity. Jugular blood DNA was extracted from healthy Cele Black sheep and Hetian sheep, and two polymorphic loci (rs399356740 and rs590747651) of \u003cem\u003eESR1\u003c/em\u003e gene in two sheep breeds were genotyped by Polymerase Chain Reaction (PCR) and Sanger sequencing, and the genetic parameters and lambing number were analyzed. The results of sequencing showed that there were three genotypes of AA, AT and TT at rs399356740 and rs590747651 loci in Cele Black sheep and Hetian sheep, and all of them showed moderate polymorphism. The dominant allele of rs590747651 locus in Cele Black sheep is T and the dominant allele of rs590747651 locus in Hetian sheep is A, which may be the reason for the difference in fecundity between the two sheep populations. The mutation site of rs590747651 was in Hardy-Weinberg equilibrium in Hetian sheep population, and the results of association analysis showed that the lambing number of AA and TT type at rs590747651 locus was significantly higher than that of AT type in Hetian sheep population, and the dominant genotype was AA type, which indicated that rs590747651 mutation could be considered as a molecular assistant marker for lambing trait in Hetian sheep. However, due to the small sample size, the rs590747651 locus had nothing to do with the litter size of Cele Black sheep. This study can be used as a potential candidate genetic marker for lambing performance in sheep breeding, and provide reference data for the study of lambing performance of sheep.\u003c/p\u003e","manuscriptTitle":"Analysis of ESR1 Gene Polymorphism and Lambing Association between Cele Black Sheep and Hetian Sheep","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2025-09-30 14:35:31","doi":"10.21203/rs.3.rs-7326024/v1","editorialEvents":[{"type":"communityComments","content":0}],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true}}],"origin":"","ownerIdentity":"ee86a65b-4e7b-401f-9c4c-a7f22c602f3c","owner":[],"postedDate":"September 30th, 2025","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"posted","subjectAreas":[],"tags":[],"updatedAt":"2025-11-20T11:24:02+00:00","versionOfRecord":[],"versionCreatedAt":"2025-09-30 14:35:31","video":"","vorDoi":"","vorDoiUrl":"","workflowStages":[]},"version":"v1","identity":"rs-7326024","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-7326024","identity":"rs-7326024","version":["v1"]},"buildId":"8U1c8b4HqxoKbykW_rLl7","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}