Detection of a new 20-bp insertion/deletion (indel) within sheep PRND gene using mathematical expectation (ME) method

Prion-related protein doppel gene (PRND), as an essential member of the mammalian prion gene family, is associated with the scrapie susceptibility as well as phenotype traits, so the genetic variation of the PRND has been highly concerned recently, including the single nucleiotide polymorphism (SNP) and insertion/deletion (indel). Therefore, the objective of present study was to examine the possible indel variants by mathematical expectation (ME) detection method as well as explore its associations with phenotype traits. A novel 20-bp indel was verified in 623 tested individuals representing 4 diversity sheep breeds. The results showed that 3 genotypes were detected and the minor allelic frequency were 0.008 (Lanzhou Fat-Tail sheep, LFTS), 0.084 (Small Tail Han sheep, STHS), 0.021(Tong sheep, TS) and 0.083 (Hu sheep, HS), respectively. Comparing with the traditional method of detecting samples one by one, the reaction times with ME method was decreased by 36.22% (STHS), 37.00% (HS), 68.67% (TS) and 83.33% (LFTS), respectively. Besides, this locus was significantly associated to cannon circumference index (P = 0.012) and trunk index (P = 0.037) in the Hu sheep breed. Notably, it was not concordance with the present result of DNA sequencing (GCTGTCCCTGCAGGGCTTCT) and dbSNPase of NCBI (NC_443194: g.46184887- 46184906delCTGCTGTCCCTGCAGGGCTT). Consequently, it was the first time to detect the new 20-bp indel of sheep PRND gene by ME strategy, which might provide a valuable theoretical basis for marker-assisted selection in sheep genetics and breeding.     

Genetic effects of PRNP gene insertion/deletion (indel) on phenotypic traits in sheep

Prion protein (PRNP) gene is well known for affecting mammal transmissible spongiform encephalopathies (TSE), and is also reported to regulate phenotypic traits (e.g. growth traits) in healthy ruminants. To identify the insertion/deletion (indel) variations of the PRNP gene and evaluate their effects on growth traits, 768 healthy individuals from five sheep breeds located in China and Mongolia were identified and analyzed. Herein, four novel indel polymorphisms, namely, Intron-1-insertion-7bp (I1-7bp), Intron-2-insertion-15bp (I2-15bp), Intron-2-insertion-19bp (I2-19bp), and 3′ UTR-insertion-7bp (3′ UTR-7bp), were found in the sheep PRNP gene. In five analyzed breeds, the minor allelic frequencies (MAF) of the above indels were in the range of 0.008 to 0.986 (I1-7bp), 0.113 to 0.336 (I2-15bp), 0.281 to 0.510 (I2-19bp), and 0.040 to 0.238 (3′ UTR-7bp). Additionally, there were 15 haplotypes and the haplotype ‘I_I2-15bp-D_3’UTR-7bp-D_I2-19bp-D_I1-7bp’ had the highest frequency, which varied from 0.464 to 0.629 in five breeds. Moreover, association analysis revealed that all novel indel polymorphisms were significantly associated with 13 different growth traits (P < 0.05). Particularly, the influences of I2-15bp on chest width (P = 0.001) in Small Tail Han sheep (ewe), 3′ UTR-7bp on chest circumference (P = 0.003) in Hu sheep, and I2-19bp on tail length (P = 0.001) in Tong sheep, were highly significant (P < 0.01). These findings may be a further step toward the detection of indel-based typing within and across sheep breeds, and of promising target loci for accelerating the progress of marker-assisted selection in sheep breeding.     

Identification of a novel 43-bp insertion in the heparan sulfate 6-O-sulfotransferase 3 (HS6ST3) gene and its associations with growth and carcass traits in chickens

Previous studies have revealed a significant association between SNPs found within the heparan sulfate 6-O-sulfotransferase 3 (HS6ST3) gene and obesity. This study identified a novel 43-bp indel polymorphism in intron 1 of HS6ST3 in 1963 chickens from nine different breeds, and three genotypes, designated II, ID and DD, were observed. The frequency of the ‘I’ (0.62–0.87) allele was higher than that of the ‘D’ (0.13–0.38) allele. A total of 777 individuals of the Gushi-Anka F₂ resource population were used for the analysis of associations according to growth traits, carcass traits, serum variables and meat quality traits. The results showed that the 43-bp indel polymorphism was significantly associated with the body weight at 4 and 6 weeks of age, chest depth at 4 and 12 weeks of age and shank girth at 12 weeks of age (P?<?0.05). In terms of the carcass traits, the indel polymorphism was significantly associated with breast muscle weight, heart weight and leg weight (P?<?0.05). These findings suggested that this indel polymorphism has the potential to become a new target for the marker-assisted selection of chicken growth and carcass traits.     

Application of mathematical expectation (ME) strategy for detecting low frequency mutations: An example for evaluating 14-bp insertion/deletion (indel) within the bovine PRNP gene

The detection method based on the mathematical expectation (ME) strategy is fast and accuracy for low frequency mutation screening in large samples. Previous studies have found that the 14-bp insertion/deletion (indel) variants of the 3′ untranslated region (3′ UTR) within bovine PRNP gene have been characterized with low frequency (≤5%) in global breeds outside China, which has not been determined in Chinese cattle breeds yet. Therefore, this study aimed to identify the 14-bp indel within PRNP gene in 5 major Chinese indigenous cattle breeds and to evaluate its associations with phenotypic traits. It was the first time to use ME strategy to detect low frequency indel polymorphisms and found that minor allele frequency was 0.038 (Qinchuan), 0.033 (Xianan), 0.013 (Nanyang), 0.003 (Jiaxian), and zero (Ji'an), respectively. Compared to the traditional detection method by which the sample was screened one by one, the reaction time by using the ME method was decreased 62.5%, 64.9%, 77.6%, 88.9% and 66.4%, respectively. In addition, the 14-bp indel was significantly associated with the growth traits in 2 cattle breeds, with the body length of Qinchuan cattle as well as the body weight and waistline of Xianan cattle. Our results have uncovered that the method based on ME strategy is rapid, reliable, and cost-effective for detecting the low frequency mutation as well as our findings provide a potential valuable theoretical basis for the marker-assisted selection (MAS) in beef cattle.     

Genetic Variant of MYLK4 Gene and its Association with Growth Traits in Chinese Cattle

As a member of MYLK family, MYLK4 gene may play a vital role in muscle development. In this study, one novel single-nucleotide polymorphism (SNP) was identified the bovine MYLK4 by sequencing pooled DNA samples (pool-Seq) and forced polymerase chain reaction-restriction fragment length polymorphism (forced PCR-RFLP) methods. Overall, we reported one mutation (SNP1) in the intron 10 region within the bovine MYLK4 gene in 559 individuals representing five main cattle breeds from China (Nanyang, NY; Qinchuan; Jiaxian, JX; Pinan cattle; and Caidamu cattle, CDM). Genotype AA and allele A were predominant in the QC, PN, and XN populations. Association analysis with growth traits in the QC breed showed that the animals with genotype GG had significantly greater chest breadth and hip width (P?<?0.05). Meanwhile, the genotype GG was strongly associated with withers height and body length than those with genotype AA (P?<?0.01 or P?<?0.05) at 12 months in the NY breed. These statistical results exhibited that the MYLK4 gene might be a potential candidate gene to improve cattle’s growth traits, and the SNP could be used as molecular markers in early marker-assisted selection (MAS) in beef cattle breeding program.     

Nucleotide variants in prion-related protein (testis-specific) gene (PRNT) and effects on Chinese and Mongolian sheep phenotypes

ABSTRACT

Studies of the ovine prion-related protein (testis-specific) gene (PRNT), including studies of genetic diversity, have highlighted its potential relationship to scrapie infection and economically important ovine traits. PRNT was previously reported to be highly polymorphic in Portuguese sheep. To characterize genetic polymorphisms in this gene in Asian sheep, a direct sequencing method was used to detect polymorphic loci in PRNT in 285 individual sheep from four Chinese and one Mongolian breeds. Seven SNP variants in PRNT were identified, including three novel variants (g.93G>A, g.162G>T, and g.190A>G) and four previously reported variants (g.17 C>T, g.112G>C, g.129C>T, and g.144A>G). In the five breeds that we analyzed, the mutation frequencies of g.190A>G in Lanzhou Fat-tail sheep (LFTS) and g.129C>T in the other four varieties were high (F>0.5). Moreover, thirteen different haplotypes that had a comparable distribution in the tested breeds were also identified; ‘C-G-G-C-A-G-A’ occurred at the highest frequency in the five sheep breeds. Additionally, we previously explored the significance of relationships between polymorphisms in PRNP or PRND and ovine growth performance. Here, we also performed correlation analysis in all tested loci. These loci polymorphisms were significantly associated with ten different growth traits (P<0.05), except for g.93G>A. Meanwhile, in contrast to a previous study, there was no significant association between the seven SNP loci analyzed and our previously reported sheep PRND or PRNP insertion/deletion mutations. Our findings may provide new insights into polymorphic variation in ovine PRNT, which may contribute to genetic improvements in economic traits that are important for sheep breeding.     

Analysis of prion protein genotypes in relation to reproduction traits in local and cosmopolitan German sheep breeds

Due to the genetic determination of susceptibility to scrapie and other forms of transmissible spongiform encephalopathy (TSE) in sheep breeding to the less susceptible prion protein (PrP) genotype ARR/ARR was advanced within EU. In 4961 ewes of nine German sheep breeds (Coburg Fox sheep, Gray Horned Heath sheep, Merinoland sheep, Rhoen sheep, German Blackheaded Mutton sheep, Shropshire, Suffolk, Texel and White East Friesian Milk sheep) representing local and cosmopolitan breeds the reproductive traits number of lambs born, dead (including abortion at the end of pregnancy, stillbirth and death during the first 56 days post natum), weaned and rearing rate at each lambing were recorded and in 1641 of these ewes the PrP genotype was determined. A linear model was used to evaluate associations between PrP genotype and reproduction traits including the effects of PrP genotype (four classes: ewes with two, one and no copy of the ARR allele and with unknown PrP genotype), breed, interaction of PrP genotype and breed, number of lambing, lambing season and stock. Significant associations were only observed between the PrP genotype and the number of dead lambs at each lambing in Shropshire and Merinoland sheep and the rearing rate at each lambing in Shropshire. These significant associations were mainly caused by differences between animals with unknown PrP genotype and animals of the other PrP classes. In conclusion, breeding for TSE resistant sheep will not lead to a reduction in economically important reproduction traits.     

Polymorphism of somatostatin gene and its association with growth traits in Chinese cattle

Somatostatins play a crucial role in the regulation of growth and development in vertebrates, especially muscle growth. We assessed the association of somatostatin gene polymorphisms with growth traits by PCR-SSCP (polymerase chain reaction-single strand conformation polymorphism) and DNA sequencing methods in 694 individuals from six Chinese cattle breeds. A novel single nucleotide polymorphism, G126A, was detected, and significant associations were found with body length, body height, hip width, heart girth, and hucklebone width index. Polymorphism of the somatostatin gene was found to be highly associated with growth traits in the Qinchuan breed at various ages. Gene frequency analysis showed significant differences among the breeds. Individuals with genotype AA had significantly lower body height, body length, hip width, and hucklebone width values compared to AG at 1.5 years old, and had significantly lower hip width, body length and hucklebone width compared to AG at 2 years old. At 2.5 years old, populations with genotype AA had significantly lower body length, hip width and hucklebone width than AG individuals, with the exception of the Luxi breed, in which two genotypes were found. The Luxi and Ximentaer crossbreed had the lowest frequency of the G allele, while the highest G allele frequencies were found in the Luxi breed.     

Polymorphism in a Microsatellite of the Acrp30 Gene and Its Association with Growth Traits in Goats

Acrp30 plays a critical role in the regulation of glucose and lipid homeostasis. In this study, polymorphism of the Acrp30 gene was detected by PCR-SSCP and DNA sequencing methods in 321 individuals from three goat breeds, and the association of Acrp30 gene polymorphism with growth traits in the three goat breeds was analyzed. A novel insert/deletion (GT)(5) microsatellite sequence was detected in the 5' flanking region of the gene. Three genotypes (AA, AB, and BB) were found in three breeds. There was moderate genetic diversity in the locus in the analyzed populations. Significant associations were observed between the genotypes of the locus and growth traits in the Boer goat population. The chest circumference of individuals with genotype BB was significantly greater than that of individuals with genotype AA.     

Genome-wide CNV analysis revealed variants associated with growth traits in African indigenous goats

Using the CaprineSNP50 data generated by the AGIN consortium, we detected common CNVs in 126 samples from four African indigenous goat breeds. A total of 30 CNVs ranging from 30,237 bp to 4,910,757 bp were identified. These CNVs were then associated with six growth traits by a linear regression analysis. Three significant associations were identified between two CNVs and two body traits after false discovery rate (FDR) correction (P < .05). One of them (CNV27) was significantly associated with both chest width and width of pin bones. It overlaps the SNX29 gene, the Gene Ontology (GO) annotations of which indicate CNV27 could be a potential functional candidate for meat production, health and reproduction traits. To our knowledge, this study is the first CNV-based association test of growth traits using SNP chip data in African meat goats.     

湖羊、东湖杂种羊POMC基因外显子3单核苷酸多态性及其与生长性状的关联分析

阿黑皮素原(Pro-opiomelanocortin, POMC)在动物采食和能量平衡调控中发挥重要作用, 文章对绵羊POMC基因外显子3进行扩增和测序, 筛选多态性位点, 并分析多态位点与湖羊和东弗里生×湖羊杂种羊生长性状的相关性。测序后发现湖羊POMC基因外显子3有2个单碱基突变(g.273 T/C和g.456 G/A), 根据273位点处发生的T/C突变, 建立PCR-RFLP分析方法, 并对162只湖羊和130只东湖杂种羊进行检测分析。结果发现, 在湖羊群体中检测到TT(0.469)、TC(0.438)和CC(0.093)3种基因型, 而在东湖杂种羊群体中仅检测到TT(0.754)和TC(0.246)两种基因型。POMC基因外显子3的273位点多态性与生长性状的相关性研究结果显示：湖羊群体中CC基因型个体的2月龄断奶重、4月龄尻高及TC基因型个体4月龄体长和管围均显著高于TT型个体(P<0.05); CC基因型个体的4月龄重、6月龄重极显著高于TT和TC基因型个体(P<0.01); CC基因型个体的4月龄体高和体长极显著高于TT型个体(P<0.01), 且显著高于TC基因型个体(P<0.05)。此外, CC型个体的管围极显著高于TT基因型个体(P<0.01)。东湖杂种羊群体中TC基因型个体的2月龄断奶重、4月龄重及4月龄体高、体长、胸深和管围都显著高于TT型个体(P<0.05), TC型个体的6月龄重极显著高于TT型个体(P<0.01)。研究结果表明, POMC基因外显子3与绵羊生长性状相关, C等位基因对体重及体尺性状的增加更有利。该结果为进一步探讨POMC基因作为绵羊生长性状的辅助选育标记奠定了基础。     

Development of a touch-down multiplex PCR method for simultaneously rapidly detecting three novel insertion/deletions (indels) within one gene: an example for goat GHR gene

Abstract

The multiplex polymerase chain reaction (PCR) method is fast and accuracy for screening several polymorphism loci in a single reaction in large samples. Therefore, this study aimed to identify three novel insertion/deletion (indel) loci in goat growth hormone receptor (GHR) gene by using multiplex PCR method and the genotypes in three indel loci are identified in 918 individuals from three Chinese goat breeds, as well as to evaluate its associations with growth traits. After validating the accuracy by taking the conventional PCR method, the concordance between these two methods was 100%. Moreover, P14 locus polymorphism was significantly associated with six growth traits of Hainan black goats. Briefly, an economic multiplex PCR method is presented to simultaneously and accurately detect three indel polymorphisms within goat GHR without the use of any special equipment, thus improving the experimental efficiency and fast obtaining the experimental result for indel genotyping.     

FecB基因在9个绵羊品种中的多态性及其与产羔数和羔羊生长发育的相关性

以绵羊BMPR-IB基因为候选基因,应用PCR-RFLP方法通过分析湖羊、夏洛来、陶赛特、萨福克、罗米丽、中国美利奴羊、中国美利奴肉用多胎品系以及陶赛特×中国美利奴羊和萨福克×中国美利奴羊杂交后代共615只个体的FecB基因多态性,以及BMPR-IB基因多态性对产羔数、体尺和体重的影响.结果表明,BMPR-IB基因在不同品种(系)绵羊中共有3种基因型(BB、B+和++),但基因型频率分布在各品种(系)间差异极显著(P<0.01).在湖羊中仅有BB基因型;在中国美利奴肉用多胎品系中BB、B+和++基因型频率分别为51%、30%和19%;而其他品种(系)羊中则仅有++基因型.对中国美利奴羊肉用多胎品系研究,发现BB和B+基因型群体平均产羔数分别为2.8和2.3,显著高于++基因型群体(1.2,P<0.01).在90日龄时,BB和B+基因型群体的体重分别为18.6±3.70 kg和18.0±3.31 kg,显著高于++基因型群体(15.6±2.22kg,P<0.05);此外,90日龄时,BB和B+基因型群体比++基因型群体胸围、胸宽较大(P<0.05);但这些差异在120日龄时消失.另外,我们还发现不同地区群体的第一胎产羔数存在明显差别.这些结果表明,BMPR-IB基因为影响绵羊产羔数的主效基因,并首次证明该基因对后代羔羊出生后生长发育具有加性效应.     

A large indel mutation of the bovine ADD1/SREBP1c gene and its effects on growth traits in some native cattle breeds from China

Adipocyte determination and differentiation-dependent factor 1/sterol regulatory element-binding protein-1c (ADD1/SREBP1c) is a major determinant of tissue differential lipogenic capacity in mammalian and avian species. The objectives of the present study were to focus on insertion-deletion polymorphism (indel) in the bovine ADD1/SREBP1c gene, and analyzing its effect on growth traits in a sample of 1035 cattle belonging to four Chinese cattle breeds. PCR-SSCP, DNA sequencing and agarose electrophoresis methods were used. The 778 bp PCR products of ADD1/SREBP1c gene exhibited three genotypes and two alleles were revealed: W and D. Frequencies of the W allele varied from 0.8651 to 1.000. The associations of the 84 bp indel mutation of ADD1/SREBP1c gene with growth traits of 265 Nanyang cows were analyzed. The animals with genotype WD had significantly higher birth weight, body weight, average daily gain than those with genotype WW at birth, 6-, 12-, 18-, and 24-month old (P < 0.05 or P < 0.01). These results suggest that the indel mutation of bovine ADD1/SREBP1c gene may influence the growth traits in cattle.     

Identification of novel SNPs of ovine <Emphasis Type="Italic">PRL</Emphasis> gene and their association with milk production traits

Prolactin (PRL) is a lactogenic hormone that plays a significant role in milk production; its depletion in sheep provokes a severe reduction of milk secretion. Thus, PRL also could be used as a positional marker gene associated with milk production and composition traits. Therefore, the purpose of the study was to identify genotype frequencies of single nucleotide polymorphisms the intron 2 in ovine PRL gene and its possible association genotypes with milk traits in dairy sheep breeds. The genetic structures of ovine PRL gene were examined by PCR-RFLP and DNA sequencing methods in three sheep populations. Four hundred and fifty blood and milk samples were used in the study, which included 150 samples from each of Sakiz, Akkaraman and Awassi ewes respectively. As a result, PRL genotype AA showed a strong association with milk yields content, whereas the animals carrying BB genotype had a higher fat percentage value in the three sheep breeds. Haplotype analysis of the obtained sequences showed the presence of 12 haplotypes in the PRL intron 2 region. In the present study, we have reported for the first time 48 SNPs of the PRL gene for intron 2 in dairy sheep breeds. These preliminary results indicate that the identified SNPs lend themselves readily for further research regarding physiological impacts such as milk production and reproductive traits in other dairy sheep populations.     

A Missense Mutation in the Rabbit Melanocortin 4 Receptor (MC4R) Gene is Associated with Finisching Weight in a Meat Rabbit Line

In this study we resequenced 1729 bp of the rabbit melanocortin 4 receptor (MC4 R) gene in 31 rabbits from different breeds/lines and identified ten polymorphisms: one was an indel and 9 were single nucleotide polymorphisms (SNPs). The indel and 5 SNPs were in the 5′-flanking region, 3 were synonymous SNPs and one was a missense mutation (c.101G>A; p.G34D), located in a conserved position of the extracellular tail of the MC4 R protein. The missense mutation was analyzed in a panel of 74 rabbits of different breeds and in 516 performance tested rabbits of a commercial paternal line under selection for growth efficiency. Association analysis indicated that rabbits with the less frequent genotype in this population (DD) had a lighter weight at 70 postnatal days than animals with genotype GD (P < 0.10) and animals with genotype GG (P < 0.05). This is the third study on candidate genes, after those on GH1 and IGF2 that reported a marker associated with finishing weight. Therefore, it seems that a candidate gene approach in rabbit based on previous information accumulated in other livestock species could be useful to identify genes explaining a fraction of variability of performance traits with potential application on rabbit breeding and selection.     

Research on association between variants and haplotypes of TRPV1 and TRPA1 genes with growth traits in three cattle breeds

The objective of this study was to examine the association of transient receptor potential vanilloid 1 (TRPV1) and transient receptor potential ankyrin 1 (TRPA1) genes polymorphisms with growth traits in three Chinese cattle breeds (Jiaxian red cattle, Qinchuan cattle and Luxi cattle). Through experiments we identified three single nucleotide polymorphisms (SNPs) in these three cattle breeds TRPV1 and TRPA1 genes using PCR-SSCP, (forced) PCR-RFLP methods. Three of these polymorphisms are all synonymous mutation which includes (NW_003104493.1: 30327 C?>?T), (NW_003104493.1: 33394 A?>?G) and (NW_003104493.1: 38471?G?>?A) are in exons. The other three polymorphisms are located at 3'UTR. Furthermore, we evaluated the haplotype frequency and the statistical analyses indicated that these SNPs of TRPV1 and TRPA1 genes were associated with bovine body height, body length, waist angle width, hucklebone width, cross ministry height, chest qingwidth (p?<?0.05) and recommendation height, cannon circumference (p?<?0.01) of Qingchuan cattle; body length, waist angle width (p?<?0.05) of Jiaxian red cattle; body weight, Body length, cannon circumference, chest circumference (p?<?0.05) and body height (p?<?0.01) of Luxi cattle. Our result confirms the polymorphisms in the TRPV1 and TRPA1 genes are associated with growth traits that may be used for marker-assisted selection (MAS) in three beef breeds programs.     

Detection of Insertions/Deletions Within <Emphasis Type="Italic">SIRT1</Emphasis>, <Emphasis Type="Italic">SIRT2</Emphasis> and <Emphasis Type="Italic">SIRT3</Emphasis> Genes and Their Associations with Body Measurement Traits in Cattle

Growth traits are complex quantitative traits controlled by numerous candidate genes, and they can be well-evaluated using body measurement traits. As the members of the nicotinamide adenine dinucleotide-dependent family of histone deacetylases, class I sirtuin genes (including SIRT1, SIRT2 and SIRT3) play crucial roles in regulating lipid metabolism, cellular growth and metabolism, suggesting that they are potential candidate genes affecting body measurement traits in animals. Hence, the objective of this work aimed to detect novel insertions/deletions (indels) of SIRT1, SIRT2 and SIRT3 genes in 955 cattle belonging to five breeds, as well as to evaluate their effects on body measurement traits. Herein, the novel 12-bp indel of SIRT1 gene, the 7-bp indel of SIRT2 gene and the 26-bp indel of SIRT3 gene were firstly reported, respectively. The association analysis indicated that the indels within SIRT1 and SIRT2 genes were significantly associated with body measurement traits such as body weight, chest circumference, height at hip cross, hip width, body height, etc. (P?<?0.05 or P?<?0.01). Therefore, based on these findings, the two novel indel variants within bovine SIRT1 and SIRT2 genes could be considered as potential molecular markers for growth traits in cattle selection practices and breeding.     

Relationship between an indel mutation within the SIRT4 gene and growth traits in Chinese cattle

Abstract

Growth traits are mainly determined by genetic factors. SIRT4, a class II sirtuin, predominantly acts as an ADP-ribosyltransferase and inhibits fatty acid oxidation. In this study, a total of 1005 cattle belonging to five indigenous Chinese breeds were used to evaluate the relationship between the potential insertions/deletions (indels) within the SIRT4 gene and growth traits. The results revealed that only one intronic variation was present, which showed Hardy–Weinberg equilibrium (p?>?0.05) in all the populations. The relationship analyses indicated that this indel was significantly associated with growth traits (p?<?0.05), implying that SIRT4 significantly affects the growth traits. Therefore, the deletion mutation within the SIRT4 gene could be considered as a molecular marker to screen for growth traits in the cattle industry.     

Polymorphism of 5′ regulatory region of ovine FSHR gene and its association with litter size in Small Tail Han sheep

Single nucleotide polymorphisms of 5?? regulatory region of follicle-stimulating hormone receptor (FSHR) gene were detected in two high prolificacy sheep breeds (Small Tail Han and Hu sheep) and two low prolificacy sheep breeds (Corriedale and Chinese Merino sheep) by polymerase chain reaction-single strand conformation polymorphism (PCR-SSCP). The results indicated that there were three genotypes (AA, AB and BB) detected by primer 1 in Hu sheep while only one genotype (AA) in other three sheep breeds, and frequencies of AA, AB and BB genotypes in Hu sheep were 0.700, 0.225 and 0.075, respectively. There were three genotypes (EE, EF and EG) detected by primer 3 in Small Tail Han sheep while only EE genotype occurred in other three sheep breeds, and frequencies of EE, EF and EG genotypes in Small Tail Han sheep were 0.775, 0.200 and 0.025, respectively. No polymorphism was detected in four sheep breeds by primer 2 and primer 4. The sequencing results showed that there were two nucleotide mutations (g. ?681T>C and g. ?629C>T) in genotype BB compared with AA for primer 1. As for primer 3, two mutations (g. ?197G>A and g. ?98T>C) in genotype EF compared with EE and two mutations (g. ?200G>A and g. ?197G>A) in genotype EG compared with EE. The heterozygous ewes with EG or EF had 0.89 (P?<?0.05) or 0.42 (P?<?0.05) lambs more than homozygous ewes (EE genotype) in Small Tail Han sheep, respectively, while there was no significant difference on litter size between EG and EF ewes.