Preservative Effects of Curcumin on Semen of Hu Sheep.

Reactive oxygen species (ROS) are important factors that lead to a decline in sperm quality during semen preservation. Excessive ROS accumulation disrupts the balance of the antioxidant system in sperm and causes lipid oxidative damage, destroying its structure and function. Curcumin is a natural plant extract that neutralizes ROS and enhances the function of endogenous antioxidant enzymes. The effect of curcumin on the preservation of sheep semen has not been reported. This study aims to determine the effects of curcumin on refrigerated sperm (4 °C) and analyze the effects of curcumin on sperm metabolism from a Chinese native sheep (Hu sheep). The results showed that adding curcumin significantly improved (p < 0.05) the viability of refrigerated sperm at an optimal concentration of 20 µmol/L, and the plasma membrane and acrosome integrity in semen were significantly improved (p < 0.05). Adding curcumin to refrigerated semen significantly increased (p < 0.05) the levels of antioxidant enzymes (T-AOC, CAT, and SOD) and significantly decreased (p < 0.05) ROS production. A total of 13,796 metabolites in sperm and 20,581 metabolites in negative groups and curcumin-supplemented groups were identified using liquid chromatography-mass spectrometry. The proportion of lipids and lipid-like molecules among all metabolites in the sperm was the highest, regardless of treatment. We identified 50 differentially expressed metabolites (DEMs) in sperm between the negative control and curcumin-treated groups. Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis revealed that DEMs were mainly enriched in the calcium signaling pathway, phospholipase D signaling pathway, sphingolipid metabolism, steroid hormone biosynthesis, 2-oxocarboxylic acid metabolism, and other metabolic pathways. The findings indicate that the addition of an appropriate concentration (20 µm/L) of curcumin to sheep semen can effectively suppress reactive oxygen species (ROS) production and extend the duration of cryopreservation (4 °C) by modulating the expression of sphingosine-1-phosphate, dehydroepiandrosterone sulfate, phytosphingosine, and other metabolites of semen. This discovery offers a novel approach to enhancing the cryogenic preservation of sheep semen.

Comprehensive analyses of 435 goat transcriptomes provides insight into male reproduction.

A systematic analysis of genes related to reproduction is crucial for obtaining a comprehensive understanding of the molecular mechanisms that underlie male reproductive traits in mammals. Here, we utilized 435 goat transcriptome datasets to unveil the testicular tissue-specific genes (TSGs), allele-specific expression (ASE) genes and their uncharacterized transcriptional features related to male goat reproduction. Results showed a total of 1790 TSGs were identified in goat testis, which was the most among all tissues. GO enrichment analyses suggested that testicular TSGs were mainly involved in spermatogenesis, multicellular organism development, spermatid development, and flagellated sperm motility. Subsequently, a total of 95 highly conserved TSGs (HCTSGs), 508 middle conserved TSGs (MCTSGs) and 42 no conserved TSGs (NCTSGs) were identified in goat testis. GO enrichment analyses suggested that the HCTSGs and MCTSGs has a more important association with male reproduction than NCTSGs. Additionally, we identified 644 ASE genes, including 88 tissue-specific ASE (TS-ASE) genes (e.g., FSIP2, TDRD9). GO enrichment analyses indicated that both ASE genes and TS-ASE genes were associated with goat male reproduction. Overall, this study revealed an extensive gene set involved in the regulation of male goat reproduction and their dynamic transcription patterns. Data reported here provide valuable insights for a further improvement of the economic benefits of goats as well as future treatments for male infertility.

Relationships between the mutations of the goat GATA binding protein 4 gene and growth traits.

Osteogenesis is a complex multilevel process regulated by multiple genes. The GATA binding protein 4 (GATA4) gene has been extensively studied for its pivotal role in bone genesis and bone differentiation. However, its relationship with the growth traits of Shaanbei white cashmere (SBWC) and Guizhou black (GB) goats remains unclear. This work aims to investigate the potential influence of genetic mutations in the GATA4 gene on the growth traits goats. Thus, two Insertion/deletion (InDel) polymorphisms (8-bp-InDel and 9-bp-InDel) were screened and detected in a total of 1161 goats (including 980 SBWC goats and 181 GB goats) using PCR and agarose gel electrophoresis. The analyses revealed that there were two genotypes (ID and DD) for these two loci. In SBWC goats, 8-bp-InDel and 9-bp-InDel loci were significantly associated with heart girth (HG) and hip width (HW). Notably, individuals with DD genotype of 8-bp-InDel locus were superior while those with DD genotype of 9-bp-InDel locus were inferior. Correlation analyses of the four combined genotypes revealed significant associations with cannon circumference (CC), body height (BH), HG and HW. This work provides a foundation for the application of molecular marker-assisted selection (MAS) in goat breeding programs. Furthermore, the findings highlight the potential of the GATA4 gene and its genetic variations as valuable indicators for selecting goats with desirable growth traits.

MiR-188-5p regulates the proliferation and differentiation of goat skeletal muscle satellite cells by targeting calcium/calmodulin dependent protein kinase II beta.

OBJECTIVE: The aim of this study was to reveal the role and regulatory mechanism of miR-188-5p in the proliferation and differentiation of goat muscle satellite cells. METHODS: Goat skeletal muscle satellite cells isolated in the pre-laboratory were used as the test material. First, the expression of miR-188-5p in goat muscle tissues at different developmental stages was detected by quantitative reverse transcription polymerase chain reaction (qRT-PCR). In addition, miR-188-5p was transfected into goat skeletal muscle satellite cells by constructing mimics and inhibitors of miR-188-5p, respectively. The changes of differentiation marker gene expression were detected by qPCR method. RESULTS: It was highly expressed in adult goat latissimus dorsi and leg muscles, goat fetal skeletal muscle, and at the differentiation stage of muscle satellite cells. Overexpression and interference of miR-188-5p showed that miR-188-5p inhibited the proliferation and promoted the differentiation of goat muscle satellite cells. Target gene prediction and dual luciferase assays showed that miR-188-5p could target the 3'untranslated region of the calcium/calmodulin dependent protein kinase II beta (CAMK2B) gene and inhibit luciferase activity. Further functional studies revealed that CAMK2B promoted the proliferation and inhibited the differentiation of goat muscle satellite cells, whereas si-CAMK2B restored the function of miR-188-5p inhibitor. CONCLUSION: These results suggest that miR-188-5p inhibits the proliferation and promotes the differentiation of goat muscle satellite cells by targeting CAMK2B. This study will provide a theoretical reference for future studies on the molecular mechanisms of skeletal muscle development in goats.

Non-Coding RNAs and Adipogenesis.

Adipogenesis is regarded as an intricate network in which multiple transcription factors and signal pathways are involved. Recently, big efforts have focused on understanding the epigenetic mechanisms and their involvement in the regulation of adipocyte development. Multiple studies investigating the regulatory role of non-coding RNAs (ncRNAs) in adipogenesis have been reported so far, especially lncRNA, miRNA, and circRNA. They regulate gene expression at multiple levels through interactions with proteins, DNA, and RNA. Exploring the mechanism of adipogenesis and developments in the field of non-coding RNA may provide a new insight to identify therapeutic targets for obesity and related diseases. Therefore, this article outlines the process of adipogenesis, and discusses updated roles and mechanisms of ncRNAs in the development of adipocytes.

Bovine HOXA11 Gene Identified from RNA-Seq: mRNA Profile Analysis and Genetic Variation Detection Using ME Method and Their Associations with Carcass Traits.

The Homeobox A11 (HOXA11) gene regulates limb skeletal development and muscle growth, thus, it was selected as a candidate gene for bovine carcass traits. In this study, we analyzed the mRNA expression level of HOXA11 in various tissues and cells, and determined the genetic variations in the HOXA11 gene, which might be used as molecular markers for cattle breeding. The mRNA expression profiles of HOXA11 in bovine different tissues showed that HOXA11 was highly expressed in both fat and muscle. The gene expression trend of HOXA11 in myoblasts and adipocytes indicated that HOXA11 might be involved in the differentiation of bovine myoblasts and adipocytes. The data in the Ensembl database showed that there are two putative insertion/deletion (InDel) polymorphisms in the bovine HOXA11 gene. The insertion site (rs515880802) was located in the upstream region (NC_037331.1: g. 68853364-68853365) and named as P1-Ins-4-bp, and the deletion site (rs517582703) was located in the intronic region (NC_037331.1: g. 68859510-68859517) and named as P2-Del-8-bp. These polymorphisms within the HOXA11 gene were identified and genotyped by PCR amplification, agarose gel electrophoresis and DNA sequencing in the 640 Shandong Black Cattle Genetic Resource (SDBCGR) population. Moreover, the mutation frequency was very low after detection, so the mathematical expectation (ME) method was used for detection. Statistical analysis demonstrated that P1-Ins-4-bp was significantly correlated with the beef shoulder (p = 0.012) and tongue root (p = 0.004). Meanwhile, P2-Del-8-bp displayed a significant correlation with the back tendon (p = 0.008), money tendon (p = 2.84 × 10-4), thick flank (p = 0.034), beef shin (p = 9.09 × 10-7), triangle thick flank (p = 0.04), triangle flank (p = 1.00 × 10-6), rump (p = 0.018) and small tenderloin (p = 0.043) in the female SDBCGR population. In summary, these outcomes may provide a new perspective for accelerating the molecular breeding of cattle through marker-assisted selection (MAS) strategies.

Investigation on mRNA expression and genetic variation within goat SMAD2 gene and its association with litter size.

The SMAD family member 2 (SMAD2), a member of the TGF-beta superfamily, executes a significant part in the oogenesis and ovulation process. A genome-wide selective sweep analysis also found SMAD2 was different in the fertility groups of Laoshan dairy goats; whether this gene was linked to litter size was unknown. Therefore, SMAD2 was chosen to study its effects on Shaanbei white cashmere goat reproduction and mRNA expression profile. Herein, the mRNA expression level of SMAD2 was firstly determined in female goat tissues, revealing significant differences in mRNA levels of different tissues (p < 0.05), including ovary tissue, indicating a potential role for SMAD2 in goat prolificacy. Then, using six pairs of primers, only one indel locus (P3-Del-12-bp) was found to be polymorphic in goat SMAD2 (n = 501). ANOVA also revealed that a P3-Del-12-bp deletion was significantly related to first-born litter size (p = 0.037). The Chi-square (χ2) test revealed that the ID genotype was significantly more prevalent in mothers with multiple lambs (p = 0.01), indicating that heterozygous individuals (ID) are more likely to produce multiple lambs. Our findings suggest that the SMAD2 gene's P3-Del-12-bp deletion could be used to improve goat breeds by assisting with litter size selection.

CircRNA Profiling Reveals an Abundant circBDP1 that Regulates Bovine Fat Development by Sponging miR-181b/miR-204 Targeting Sirt1/TRARG1.

The proliferation and differentiation of preadipocytes is an important factor determining bovine fat development, which is closely related to the feed conversion ratio, carcass traits, and beef quality. The purpose of this study was to identify the effects of candidate circRNA and miRNA on the proliferation and differentiation of bovine preadipocytes in order to provide basic materials for molecular breeding in cattle. circRNA sequencing was performed on bovine adipocyte samples at different differentiation time points, and a total of 1830 differentially expressed circRNAs were identified. Among them, circBDP1, derived from the bovine BDP1 gene, has potential binding sites for miR-204 (known as a regulator of bovine fat development) and miR-181b, which gives us a hint that circBDP1 may regulate bovine fat development by adsorbing miR-204 and miR-181b. Here, our results revealed that circBDP1 overexpression promoted the proliferation and differentiation of bovine preadipocytes. The miRNA profile of bovine adipocytes at different differentiation time points was also analyzed using the small RNA sequencing method, and a total of 89 differentially expressed miRNAs were identified, including miR-204 and miR-181b. As expected, dual-luciferase reporter results showed that circBDP1 competitively adsorbed miR-181b and miR-204. Overexpression and interference of miR-181b in bovine preadipocytes and 3T3-L1 showed that miR-181b promoted the proliferation and differentiation of preadipocytes. Further results displayed that miR-181b and miR-204 simultaneously targeted the SIRT1 gene, and miR-204 also targeted the 3' UTR region of the TRARG1 gene. In summary, this study found that miR-181b and miR-204 were involved in fat development by targeting SIRT1 and TRARG1. The results of this study will lay a foundation for the research of fat development and beef cattle industry.

Goat SMAD family member 1 (SMAD1): mRNA expression, genetic variants, and their associations with litter size.

SMAD family member 1 (SMAD1) is phosphorylated and activated by the BMP receptors, which help regulate ovulation rate, cell growth, apoptosis, and development. Previously, the genome-wide association study revealed that it has been associated with fecundity in sheep. However, its effect on litter size has not been investigated in goats. Therefore, this study aimed to determine the level of SMAD1 mRNA expression in various tissues and to identify its polymorphisms and their association with litter size in Shaanbei white cashmere goat (SBWC). As a result, RT-qPCR analysis showed that SMAD1 was expressed in various tissues in female SBWC goats, including the ovary (P < 0.05). Importantly, the mRNA expression level in the ovaries of mothers of multi-lambs had a higher level than the mothers of single lambs (P < 0.05). Moreover, two InDels (18-bp and 7-bp) in intron 1 of SMAD1 were polymorphic among ten potential loci. Both 18-bp and 7-bp InDels were significantly correlated with litter size (P = 0.014) and (P = 0.0001), respectively. As shown by the chi-squared test, genotypic distributions of 18-bp and 7-bp were significantly distinct between single-lamb (P = 0.02) and multi-lamb mothers (P = 0.002). Our findings confirm that two InDels in SMAD1 were significantly associated with litter size and suggest that they could be used to improve fertility traits in goat breeding strategies.

Genetic Variations within the Bovine CRY2 Gene Are Significantly Associated with Carcass Traits.

As an important part of the circadian rhythm, the circadian regulation factor 2 of cryptochrome (CRY2), regulates many physiological functions. Previous studies have reported that CRY2 is involved in growth and development. However, the relationship between CRY2 gene polymorphism and cattle carcass traits remains unclear. The aim of this study was to detect the possible variations of the CRY2 gene and elucidate the association between the CRY2 gene and carcass traits in the Shandong Black Cattle Genetic Resource (SDBCGR) population (n = 705). We identified a 24-bp deletion variation (CRY2-P6) and a 6-bp insertion variation (CRY2-P7) in the bovine CRY2 gene. The frequency of the homozygous II genotype is higher than the heterozygous ID genotype in both two loci. In addition, CRY2-P6 was consistent with HWE (p > 0.05). Importantly, the CRY2-P6 variant was significantly associated with 12 carcass traits, including gross weight, ribeye, high rib, thick flank, etc. and the II was the dominant genotype. The CRY2-P7 site was also significantly correlated with five traits (gross weight, beef-tongue, etc.). Collectively, these outcomes indicated that the two Indel loci in the CRY2 gene could be used for marker-assisted selection of cattle carcass traits.

Screening of Bovine Tissue-Specific Expressed Genes and Identification of Genetic Variation Within an Adipose Tissue-Specific lncRNA Gene.

Global classification of bovine genes is important for studies of biology and tissue-specific gene editing. Herein, we classified the tissue-specific expressed genes and uncovered an important variation in the promoter region of an adipose tissue-specific lncRNA gene. Statistical analysis demonstrated that the number of genes specifically expressed in the brain was the highest, while it was lowest in the adipose tissues. A total of 1,575 genes were found to be significantly higher expressed in adipose tissues. Bioinformatic analysis and qRT-PCR were used to uncover the expression profiles of the 23 adipose tissue-specific and highly expressed genes in 8 tissues. The results showed that most of the 23 genes have higher expression level in adipose tissue. Besides, we detected a 12 bp insertion/deletion (indel) variation (rs720343880) in the promoter region of an adipose tissue-specific lncRNA gene (LOC100847835). The different genotypes of this variation were associated with carcass traits of cattle. Therefore, the outcomes of the present study can be used as a starting point to explore the development of cattle organs and tissues, as well as to improve the quality of cattle products.

CircRNA Profiling Reveals CircPPARγ Modulates Adipogenic Differentiation via Sponging miR-92a-3p.

Adipogenesis describes the proliferation, differentiation, and apoptosis of mature adipocytes from primary adipocytes and is regulated by post-transcriptional modifications. Circular RNAs (circRNAs) play critical roles in mammalian development and physiology. However, the circRNA-mediated regulation of adipogenesis remains poorly understood. We profiled circRNA expression during bovine primary adipogenesis, detecting 16 circRNA candidates, including circPPARγ, which was abundant in the adipose tissue. Overexpression (overexpression plasmids) and interference (small interfering RNAs) with circPPARγ in bovine primary adipocytes, and proliferation, differentiation, and apoptosis were analyzed using EdU (5-ethynyl-2'-deoxyuridine) cell proliferation, cell counting kit-8, flow cytometry, TdT-mediated dUTP nick-end labeling apoptosis assay, Oil Red O staining, quantitative real-time PCR, and western blotting assays, which showed that circPPARγ facilitates adipocyte differentiation and inhibits proliferation and apoptosis. Dual-luciferase reporter assay and RNA immunoprecipitation assays indicated that circPPARγ binds miR-92a-3p and YinYang 1 (YY1). A novel regulatory pathway regulating adipogenesis and adipose deposition was revealed.

Corrigendum: Novel lncRNA lncFAM200B: Molecular Characteristics and Effects of Genetic Variants on Promoter Activity and Cattle Body Measurement Traits.

[This corrects the article DOI: 10.3389/fgene.2019.00968.].

Bovine TMEM95 gene: Polymorphisms detecting in five Chinese indigenous cattle breeds and their association with growth traits

BACKGROUND: Transmembrane protein 95 (TMEM95) plays a role in male fertility. Previous studies showed that genes with a significant impact on reproductive traits can also affect the growth traits of livestock. Thus, we speculated that the genetic variation of TMEM95 gene may have effects on growth traits of cattle. RESULTS: Two SNPs were genotyped. The rs136174626 and rs41904693 were in the intron 4 and 30 -untranslated region, respectively. The linkage disequilibrium analysis illustrated that these two loci were not linked. The rs136174626 was associated with six growth traits of Nanyang cattle, four traits of Luxi cattle, and three traits of Ji'an cattle. For rs41904693 locus, the GG individuals had greater body height and abdominal girth in Ji' an cattle than TT and TG individuals. In Jinnan cattle, GG and TT individuals had greater body height, height at hip cross, body length, and heart girth than TG individuals. The potential splice site prediction results suggest that the rs136174626 may influence the splicing efficiency of TMEM95, and the miRNA binding site prediction results showed that the rs41904693 may influence the expression of TMEM95 by affecting the binding efficiency of Bta-miR-1584 and TMEM95 30 -UTR. CONCLUSIONS: The findings of the study suggested that the two SNPs in TMEM95 could be a reliable basis for molecular breeding in cattle.

Identification of novel alternative splicing of bovine lncRNA lncFAM200B and its effects on preadipocyte proliferation.

Adipogenesis is closely related to human health, livestock growth, and meat quality. A previous study identified that bovine lncFAM200B promoter has high activity in 3T3-L1 mice preadipocytes. Thus, lncFAM200B was a candidate gene for regulating adipogenesis. This study aimed to uncover the role of lncFAM200B in bovine adipogenesis and identify novel genetic variations within the bovine lncFAM200B gene. An expression analysis found that lncFAM200B was expressed higher in fat than that in muscle, but the difference was not related to the total methylation level of the promoter active region. Moreover, the expression of lncFAM200B exhibited a significant positive correlation with the expression of C/EBPa during bovine adipocyte differentiation. To uncover the function of lncFAM200B, the full-length lncFAM200B was cloned, and four kinds of transcript variants were found. Protein-coding potential prediction and prokaryotic expression system analysis showed that these four transcript variants were noncoding RNAs. The quantitative reverse-transcription polymerase chain reaction and 5-ethynyl-2'-deoxyuridine assay showed that the transcript variants decreased the messenger RNA expression of Cyclin D1 and inhibited the proliferation of bovine preadipocytes. Considering the important role of lncFAM200B in adipogenesis, we identified genetic variations in lncFAM200B. Three single-nucleotide polymorphisms (SNPs) were revealed, and two of them (SNP1 and SNP3) were associated with Nanyang cattle body measurement traits. In conclusion, this study found that bovine lncFAM200B inhibited preadipocyte proliferation, and two genetic variations of lncFAM200B could be used in cattle breeding.

Goat sperm associated antigen 17 protein gene (SPAG17): Small and large fragment genetic variation detection, association analysis, and mRNA expression in gonads.

Transcriptome sequencing analyses have suggested that sperm associated antigen 17 protein gene (SPAG17) may play important regulating roles in litter size. In this study, the expression profiles and genetic variations of the SPAG17 were studied in Shaanbei White Cashmere (SBWC) goats (n=1567). SPAG17 was highly expressed in testis and ovary of SBWC goats. At different developmental stages, it also continued to be highly expressed in testis. In addition, two variations of SPAG17, one indel locus and one copy number variation locus, were significantly associated with first-born litter size. Joint analysis results suggested that two polymorphic loci of the SPAG17 gene may regulate host gene expression in goat ovary and testis. Overall, the results indicated the important role of SPAG17 in the reproductive process of goats.

Effects of genetic variations within goat PITX2 gene on growth traits and mRNA expression.

Paired-like homeodomain 2 (PITX2), a key gene in hypothalamic-pituitary-adrena axis, influences animal growth and development. The objective of this study was to identify the association of the functional genetic variations within goat PITX2 gene with growth traits and mRNA expression levels. According to the reported single nucleotide polymorphisms (SNPs) information in Guanzhong dairy goat (GZDG), we identified genotypes of the known SNPs in Hainan black goat (HNBG). Association analysis uncovered that the SNPs of AC_000163: g.18117T > C, g.18161C > G and g.18353T > C loci were significantly associated with several growth traits (e.g., body height and body length) in HNBG and GZDG breeds. According to the quantitative real-time PCR assay, ß-Actin and ribosomal protein L19 (RPL19) were the most stable expressed housekeeping genes in heart and skeletal muscle, respectively; meanwhile, glyceraldehyde-3-phosphate dehydrogenase (GAPDH) was the most stable expressed housekeeping gene in the other tissues. Based on the best housekeeping gene of varied tissues, we found the different genotypes of above loci were significantly associated with PITX2 mRNA expression in heart, muscle and small intestine. Briefly, the genetic variants of goat PITX2 associating with mRNA expression affected growth traits significantly, which would benefit for goat breeding in the future.

DNA methylation pattern of the goat PITX1 gene and its effects on milk performance.

Paired-like homeodomain transcription factor 1 (PITX1) is a pivotal gene in the hypothalamic-pituitary-adrenal axis, which is a well-known pathway affecting lactation performance. The aim of this study was to analyze the DNA methylation profile of the PITX1 gene and its relevance to milk performance in Xinong Saanen dairy goats; thus, potential epigenetic markers of lactation performance were identified. A total of 267 goat blood samples were divided into "low" and "high" groups according to two milk traits: the average milk yield (AMY) and the average milk density (AMD). One CpG island in the 3 ' -flanking region of the PITX1 gene was identified as being related to milk performance. Fisher's exact test demonstrated that the methylation rates of the overall CpG island and the 3rd and 12th CpG-dinucleotide loci in the blood were significantly associated with the AMY, and the overall methylation rate of the high AMY group was relative hypomethylation compared with the low AMY group. The overall methylation rates of this CpG island in mammary gland tissue from dry and lactation periods again exhibited a significant difference: the lactation period showed relative hypomethylation compared with the dry period. Bioinformatic transcription factor binding site prediction identified some lactation performance related transcription factors in this CpG island, such as CTCF, STAT, SMAD, CDEF, SP1, and KLFS. Briefly, overall methylation changes of the CpG island in the PITX1 gene are relevant to lactation performance, which will be valuable for future studies and epigenetic marker-assisted selection (eMAS) in the breeding of goats with respect to lactation performance.

lnc9141-a and -b Play a Different Role in Bovine Myoblast Proliferation, Apoptosis, and Differentiation.

Previously, our transcriptome sequencing revealed that lnc9141 was differentially expressed in muscles of fetal bovine, calf, and adult bovine, which is considered to provide the basis for raising the muscle mass. In this study, we identified lnc9141 characters. lnc9141 has different transcription start sites and 3' alternative splicing sites of exon 1, producing lnc9141-a and lnc9141-b transcripts that were highly expressed in the heart and lung. Moreover, neither lnc9141-a nor lnc9141-b had the ability to encode proteins. The functions of lnc9141-a and lnc9141-b were explored by cell cycle, 5-ethynyl-2'-deoxyuridine (EdU), and 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. The results showed that lnc9141-a or lnc9141-b overexpression decreased the number of myoblasts in the S phase and increased the proportion of cells in the G0/G1 phase. Furthermore, overexpressing lnc9141-a and lnc9141-b respectively downregulated the expression of Cyclin D1. However, lnc9141-a or lnc9141-b interference was found to increase the number of S-phase myoblasts, and upregulate Cyclin D1 and Cyclin E expression. Through Annexin V-FITC/propidium iodide (PI) double staining and the expression of apoptosis marker genes (Bax, Bcl2, and Caspase-3), it was found that lnc9141-b could regulate the expression of Bax gene. Meantime, high expression of lnc9141-b could decrease MyHC expression. In addition, the intergenic region between lnc9141 and IRX5 was 2.3 kb, with a head-to-head orientation. The study also revealed the core regions of the lnc9141 and IRX5 promoter. Our study demonstrated that both lnc9141-a and -b expression inhibited bovine myoblast proliferation. However, lnc9141-b regulated Bax and MyHC expression. The regulatory mechanism of lnc9141-a and lnc9141-b needs to be further explored.

Genetic Effects of Single Nucleotide Polymorphisms in the Goat GDF9 Gene on Prolificacy: True or False Positive?

Goat reproductive traits are complex quantitative traits controlled by polygenes and multipoint. To date, some high-fertility candidate genes in livestock have been unearthed and the growth differentiation factor 9 (GDF9) gene is one of them, which plays a crucial role in early folliculogenesis. According to the relevant previous studies and the National Center for Biotechnology Information Search database (NCBI), a total of 45 single nucleotide polymorphisms (SNPs) have been detected in the goat GDF9 gene, but which one or which ones have important effects on goat fecundity is still uncertain. Hence, in order to find effective molecular markers for goat genetic breeding and accelerate the goat improvement, this study summarized and classified the above 45 SNPs into four kinds, as well as compared and analyzed the same SNP effects and the different SNPs linkage effects on the reproductive traits in different goat breeds. Since there were many SNPs in the goat GDF9 gene, only 15 SNPs have been identified in more than 30 goat breeds worldwide and they showed different effects on the litter size. Therefore, this study mainly chose these 15 SNPs and discussed their relationship with goat productivity. Results showed that three non-synonymous SNPs A240V, Q320P, and V397I and three synonymous ones L61L, N121N, and L141L played a "true" role in the litter size trait in many goat breeds around the world. However, the regulatory mechanisms still need further research. These results provide an effective tool for follow-up research developing the goat molecular breeding strategies and improving the goat reproductive traits.