Preliminary studies on the molecular mechanism of intramuscular fat deposition in the longest dorsal muscle of sheep.

BACKGROUND: Intramuscular fat content is an important index reflecting the quality of mutton, which directly affects the flavor and tenderness of mutton. Livestock and poultry intramuscular fat content is influenced by genetics, nutritional level, and environmental factors. Key regulatory factors play a crucial role in intramuscular fat deposition. However, there is a limited amount of research on the identification and function of key genes involved in intramuscular fat content deposition specifically in sheep. RESULTS: Histological differences in the longest dorsal muscle of the small-tailed frigid sheep increased in diameter and decreased in several muscle fibers with increasing monthly age; The intramuscular fat content of the longest dorsal muscle of the small-tailed cold sheep varied with age, with a minimum of 1 month of age, a maximum of 6 months of age, and a minimum of 12 months of age. Transcriptomic sequencing and bioinformatics analysis revealed a large number of differential genes in the longest dorsal muscles of little-tailed billy goats of different months of age, which were enriched in multiple GO entries and KEGG pathways. Among them, the pathway associated with intramuscular fat was the AMPK signaling pathway, and the related genes were PPARGC1A and ADIPOQ; Immunohistochemical studies showed that PPARGC1A and ADIPOQ proteins were expressed in connective tissues, cell membranes, and, to a lesser extent, the cytoplasm of the longest dorsal muscle of the little-tailed frigid sheep; Real-time PCR and Western Blot validation showed that PPARGC1A and ADIPOQ were both expressed in the longest dorsal muscle of the little-tailed frigid sheep at different ages, and there were age differences in the amount of expression. The ADIPOQ gene was negatively correlated with the intramuscular fat content of the longest dorsal muscle, and the PPARGC1A gene was positively correlated with the intramuscular fat content of the longest dorsal muscle; As inferred from the above results, the ADIPOQ gene was negatively correlated with the intramuscular fat content of the longest dorsal muscle (r = -0.793, P < 0.05); and the PPARGC1A gene was positively correlated with the intramuscular fat content of the longest dorsal muscle r = 0.923, P < 0.05). CONCLUSIONS: Based on the above results, it can be inferred that the ADIPOQ gene is negatively correlated with the intramuscular fat content of the longest back muscle (r = -0.793, P < 0.05); the PPARGC1A gene is positively correlated with the intramuscular fat content of the longest back muscle (r = 0.923, P < 0.05).

Genome-wide association research on the reproductive traits of Qianhua Mutton Merino sheep.

Objective: Qianhua Mutton Merino sheep is a new breed of meat wool sheep cultivated independently in China. In 2018, it was approved by the state and brought into the national list of livestock and poultry genetic resources. Qianhua Mutton Merino sheep has the common characteristics of typical meat livestock varieties with rapid growth and development in the early stage and high meat production performance.The objective of this research is to investigate the Genome-wide association of the reproductive traits of Qianhua Mutton Merino sheep. Methods: Qianhua Mutton Merino sheep from the breeding core group were selected as the research object, GWAS was conducted on genes associated with the reproductive traits (singleton or twins, birth weight, age [in days] for sexual maturity, weaning weight, and daily gain from birth to weaning) of Qianhua mutton merino. Results: Our study findings showed that 151 loci of SNPs were detected, among which 3 SNPs related to birth weight and weaning weight occupied a significant portion of the wide genome. The candidate genes preliminarily obtained were SYNE1, SLC12A4, BMP2K, CAMK2D, IMMP2L, DMD, and BCL2. Conclusion: 151 SNP loci were detected for five traits related to reproduction (including singleton or twins, birth weight, age [in days] at sexual maturity, weaning weight, and daily weight gain from birth to weaning). The functions of these candidate genes were mainly enriched in nucleotide metabolism, metal ion binding, oxytocin signaling pathway, and neurotrophin signaling pathway.

Fitting of Growth Curves and Estimation of Genetic Relationship between Growth Parameters of Qianhua Mutton Merino.

Qianhua Mutton Merino is a dual-purpose (meat and wool) breed of sheep that has been newly developed in China. In this study, we assessed the growth and development of the Qianhua Mutton Merino sheep breed under house feeding conditions by measuring the body weight and chest circumference of 2300 rams and ewes of this breed aged 0-24 months. Based on the fitting results of three nonlinear growth models, namely Logistic, Gompertz, and von Bertalanffy, in Qianhua Mutton Merino, we selected the von Bertalanffy model because of its highest fitting degree among all models (R2 > 0.977). The significant analysis of the combined fixation of each sheep body's weight and bust took place (A: mature body weight, B: adjustment parameter, K: instant relative growth rate). The results revealed that parameters A, B, and K of body weight and chest circumference have high heritability and thus could be used as target traits for genetic improvement. Moreover, the correlation strength among A, B, and K suggested that these parameters can be used as a reference to adjust the genetic parameters in the growth model to genetically improve the body size of Qianhua Mutton Merino during breeding.

MiR-21 regulated hair follicle cycle development in Cashmere goats by targeting FGF18 and SMAD7.

Increasing Cashmere production can add value because it is the primary product of Cashmere goats. Recent years, peoples find miRNAs are crucial in regulating the development of hair follicle. Following Solexa sequencing, many miRNAs were distinguishingly expressed in telogen skin samples of goats and sheep in earlier study. But the method through which miR-21 controls the growth of hair follicles is still ambiguous. Bioinformatics analysis was used to predict the target genes of miR-21. The mRNA level of miR-21 in telogen Cashmere goat skins was higher than in anagen, according to the results of qRT-PCR, and the target genes expressed similarly with miR-21. Western blot showed similar trend, the protein expression of FGF18 and SMAD7 were lower in anagen samples. The Dual-Luciferase reporter assay confirmed miRNA-21's relationship with its target gene, and the consequences indicated found FGF18 and SMAD7 have positive correlations with miR-21. Western blot and qRT-PCR distinguished the expression of protein and mRNA in miR-21 and its target genes. According to the consequence, we found that target genes expression was increased by miR-21 in HaCaT cells. This study identified that miR-21 might take part in the development of Cashmere goat's hair follicles by targeting FGF18 and SMAD7.

A 1.1 Mb duplication CNV on chromosome 17 contributes to skeletal muscle development in Boer goats.

The Boer goat is one of the top meat breeds in modern animal husbandry and has attracted widespread attention for its unique growth performance. However, the genetic basis of muscle development in the Boer goat remains obscure. In this study, we identified specific structural variants in the Boer goat based on genome-wide selection signals and analyzed the basis of the molecular heredity of related candidate genes in muscle development. A total of 9 959 autosomal copy number variations (CNVs) were identified through selection signal analysis in 127 goat genomes. Specifically, we confirmed that the highest signal CNV (HSV) was a chromosomal arrangement containing an approximately 1.11 Mb (CHIR17: 60062304-61171840 bp) duplicated fragment inserted in reverse orientation and a 5 362 bp deleted region (CHIR17:60145940-60151302 bp) with overlapping genes (e.g., ARHGAP10, NR3C2, EDNRA, PRMT9, and TMEM184C). The homozygous duplicated HSV genotype (+/+) was found in 96% of Boer goats but was not detected in Eurasian goats and was only detected in 4% of indigenous African goats. The expression network of three candidate genes ( ARHGAP10, NR3C2, and EDNRA) regulating dose transcription was constructed by RNA sequencing. Results indicated that these genes were involved in the proliferation and differentiation of skeletal muscle satellite cells (SMSCs) and their overexpression significantly increased the expression of SAA3. The HSV of the Boer goat contributed to superior skeletal muscle growth via the dose effects of overlapping genes.

MiR-125b regulates the differentiation of hair follicles in Fine-wool Sheep and Cashmere goats by targeting MXD4 and FGFR2.

With the development of miRNAs identification technology, more and more miRNAs have been discovered, and the role of miRNAs in the development of animal hair follicles has become a focus of research on hair-producing animals. In the previous experiment, compare the microRNA (miRNA) trancriptomes of goats and sheep skin using Solexa sequencing and differentially expressed miR-125b was screened. However, the mechanism of miR-125b regulating hair follicle development is not clear. Therefore, in the present study, the expression of miR-125b, MXD4 and FGFR2 in skin tissue of Fine-wool Sheep and Cashmere goats and HEK-293T cells was examined by qPCR and Western blot. Furthermore, the correlation between miR-125b and the predicted target gene (MXD4, FGFR2) was verified using the Dual-luciferase Reporter assay. We demonstrated that the expression of MXD4 and FGFR2 in Cashmere goats was significantly higher than that of Fine-wool Sheep, and the expression was opposite to that of miR-125b. miR-125b can down-regulate the levels of MXD4 and FGFR2. Dual-luciferase reporter gene assay showed that miR-125b could bind to the 3'-UTR region of target genes FGFR2 and MXD4, suggesting that MXD4 and FGFR2 were target genes of miR-125b. This study has shown that the growth and development of hair follicles in skin tissue of Fine-wool Sheep and Cashmere goats from the new regulatory levels of miRNAs, and clarified the mechanism of miR-125b and its target genes in the development of hair follicles in the skin.

Expression of miRNA-1-3p and its target gene in hair follicle cycle development of Liaoning Cashmere goat.

MicroRNA exerts an important regulatory role in almost all the biological process, including hair follicle development in Liaoning Cashmere goat. In order to improve the Cashmere performance of goat, the regulatory role of microRNA in hair follicle cycle has drawn hotspot attention. However, the molecular mechanisms of miRNA-1-3p involved in hair follicle development are poorly understood. In this study, we found that miRNA-1-3p was less expressed in anagen stage of hair follicle cycle of Cashmere goat than that in telogen stage by using RT-qPCR and immunoblotting analysis, in contrast to the expression pattern of FGF14. The Dual-Luciferase reporter assay was employed to verify the relationship between miRNA-1-3p and FGF14. The results showed that miRNA-1-3p specifically binds to the 3'UTR of FGF14 mRNA, and FGF14 is the target gene of miR-1-3p. In conclusion, this study shows that miRNA-1-3p may regulate hair follicle development in Liaoning Cashmere goats by targeting FGF14.

MiR-19 3b regulated the formation of coat colors by targeting WNT10A and GNAI2 in Cashmere goats.

MiRNAs as a series of small noncoding RNAs that play a crucial part in regulating coat color and hair follicle development. In the previous Solexa sequencing experiments, there were many miRNAs expressed differentially in alpacas with different coat color, including miR-193b.But the mechanism of miR-193b in mammalian pigmentation is still unknown. In this study, bioinformatics analysis showed that WNT10A and GNAI2 might be the target genes of miR-193b. qRT-PCR showed the expression of miR-193b in white Cashmere goats' skins was obviously lower than that in browns, and the expression of WNT10A and GNAI2 were similar with miR-193b. The protein levels of WNT10A and GNAI2 indicated the same findings. Furthermore, the expression of WNT10A and GNAI2 in keratinocytes were analyzed from mRNA and protein levels, the results manifested that the group of overexpression of miR-193b in HaCaT cells increased the expressions of target genes, and miR-193b inhibition group reduced expressions. Luciferase report assays confirmed that the targeting relationship between miR-193b and target genes (WNT10A and GNAI2), the results showed that miR-193b was positively correlated with target genes. These experimental data showed that miR-193b might participate in adjustment of coat color in skin tissue of Cashmere goat by targeting WNT10A and GNAI2.

Research on meat quality of Qianhua Mutton Merino sheep and Small-tail Han sheep.

In this study, we analyzed the meat quality of Qianhua Mutton Merino sheep (QHMM). QHMM and Small-tail Han sheep (STH) were selected as the test animals. After slaughtering, the longissimus dorsi muscles were tested to determine the nutrient composition, content of amino acid, fatty acid, and muscle fiber diameter. According to the nutrient test result, the protein content of QHMM was higher than that of STH. However, the fat content was significantly lower (P < 0.05) than STH. Additionally, total amino acid content, total essential amino acid content, total half-essential amino acid content, total non-essential amino acid content, and flavor amino acid content of QHMM were significantly higher (P < 0.05) than those of STH. Moreover, the contents of C22:1 n9, C18:2 n6, C18:3 n6, C22:6 n6, and C10:0 of the muscle in QHMM were significantly higher (P < 0.05) than those of STH. Furthermore, the essential amino acid score, the total unsaturated-to-saturated fatty acid ratio, and the polyunsaturated-to-saturated fatty acid ratio of QHMM were greater than those of STH. Additionally, the muscle fiber diameter of QHMM was considerably lower (P < 0.01) than that of STH. In conclusion, the meat quality of QHMM was better than that of STH.

Stepwise Method and Factor Scoring in Multiple Regression Analysis of Cashmere Production in Liaoning Cashmere Goats.

Liaoning cashmere goat is a well-known local cashmere goat breed in China and even in the world. It is famous for producing cashmere with superior quality and high yield. Cashmere yield, body measurements, and body weight are the primary indicators of cashmere goat breeding, but the correlation between them is not yet clear. Therefore, this study investigated the relationship between certain body measurements, body weight, and cashmere yield in Liaoning cashmere goats using stepwise and factor score analyses in a multiple regression analysis. For this purpose, the body measurements (body slanting length (BSL), body height (BH), chest circumference (CC), pipe circumference (PC), chest depth (CD), chest width (CW), hip breadth (HB), body weight (BW) and cashmere yield (CY)) of 200 (2-year-old) Liaoning cashmere goats were collected. Stepwise analysis of the results showed that body weight had the greatest direct effect on cashmere yield, followed by hip breadth, while chest circumference mainly affected cashmere yield indirectly. The results of factor score analysis showed that the independent variable can be represented by two factors, which explained 49.596% and 12.095% of the total variance, respectively. The factor scores used in the regression analysis explained 75.8% of the total variance in Liaoning cashmere yield. The above studies show that the growth traits of Liaoning cashmere goats are closely related to the cashmere yield. Growth traits should be considered important factors in breed selection, germplasm identification, and rearing.

Exogenous Melatonin Directly and Indirectly Influences Sheep Oocytes.

Understanding whether and how melatonin (MT) may impact sheep oocyte development competence is central to our ability to predict how sheep oocytes will respond to artificially regulated estrus. Implanting MT can make sheep enter estrus during the non-breeding season. One study found that the blastocyst rate increased under MT treatment, while another found that the blastocyst rate decreased. Therefore, we conducted a meta-analysis of MT directly and indirectly influencing sheep oocytes. A total of 433 articles were collected from which 20 articles and 34 treatments were finally selected. A method for estimating the default value was established for the litter size analysis. We found that exogenous MT add into in vitro maturation medium was positively related to the blastocyst rate in the lab. However, subcutaneous implanting MT did not affect the in vivo ovulation rate, fertilization rate, blastocyst rate, or pregnancy rate at farm. MT did not affect the in vitro cleavage rate. However, MT improved the in vivo cleavage rate. We hypothesized that implanted MT could increase the concentration of MT in oviduct fluid in vivo, and also that in vitro MT could increase the early cleavage rate of sheep zygotes without affecting the total cleavage rate. In the analysis of oocyte apoptosis caused by injury, the results suggested that pyroptosis would be more suitable for further research. MT produces responses in all body organs, and thus implanting of MT during non-breeding seasons should consider the effect on animal welfare.

MicroRNA-200a regulates skin pigmentation by targeting WNT5A and FZD4 in Cashmere goats.

MicroRNAs are small, non-coding RNAs that regulate the expression of target genes. Previous research has demonstrated that microRNA-200a regulates cell apoptosis, tumor progression, and autoimmune disease. Preliminary studies found that microRNA-200a was differently expressed in the skin of Cashmere goats of various coat colors. However, the role of microRNA-200a in skin pigmentation remained poorly understood. In the current study, we investigated the effect of microRNA-200a on pigmentation in Cashmere goats. The expression of target genes was detected by real-time quantitative PCR, western blot, and immunohistochemistry staining both in vivo and in vitro. Luciferase reporter assays were used to demonstrate the relationship between microRNA-200a and its target genes Wnt family member 5A and frizzled class receptor 4 (WNT5A and FZD4) in HEK293T cells. BALB/c mice were injected with antagomiR-200a to detect melanin content and the expression of microRNA-200a and its target genes. The results demonstrated that the expression of microRNA-200a was significantly higher in brown tissue. Luciferase reporter assays confirmed that microRNA-200a targeted WNT5A and FZD4. The expression of WNT5A and FZD4 in the skin of brown Cashmere goats was significantly lower than that in white Cashmere goats by the detection of mRNA and protein levels. Overexpression/inhibition of microRNA-200a in keratinocytes decreased/increased the mRNA and protein expression of WNT5A and FZD4, respectively. In addition, the expression of WNT5A and FZD4 increased in the skin of BALB/c mice injected with antagomiR-200a, but the melanin content decreased. In summary, this study indicated that microRNA-200a regulates skin pigmentation by targeting WNT5A and FZD4 in Cashmere goats.

Expression of miRNA-203 and its target gene in hair follicle cycle development of Cashmere goat.

MicroRNA plays an important regulatory role in the development of all organisms, including hair follicle development. In order to improve domestic cashmere yield, the role of miRNA in hair follicle cycle has become a research hotspot. However, the molecular and cellular mechanisms by which miRNA-203 regulates hair follicle development are not completely understood. In this study, we found that the relevant target genes of miRNA-203 (DDOST and NAE1) were less expressed in telogen by qPCR and Immunoblotting analysis, contrary to the expression mode of miRNA-203. The Dual-Luciferase reporter assay was used to verify the correlation between miRNA-203 and its target gene expression. The results showed that miRNA-203 specifically binds to the 3 'UTR of DDOST and NAE1, and the expression of miRNA-203 significantly down-regulates the expression of DDOST and NAE1 mRNA and protein. Therefore, this study demonstrates that miRNA-203 may regulate hair follicle development in Cashmere goats by targeting DDOST and NAE1.

MiR-27a regulates WNT3A and KITLG expression in Cashmere goats with different coat colors.

MicroRNAs(miRNAs) regulate and control gene expression at the post-transcriptional level by base pairing with its target gene 3'UTR, resulting in degradation of the target mRNA or inhibition of its translation. The previous high-throughput sequencing results indicated that miR-27a was involved in coat color regulation. However, the mechanism of action is not still illuminated. In this research, using western blotting and real-time quantitative polymerase chain reaction(qRT-PCR), the expression of miR-27a, WNT3A and KITLG were examined in the skin of Cashmere goats with white and brown, and human embryonic kidney 293 T cells (HEK-293T cells) which over-express miR-27a. Targeting relationship between miR-27a and WNT3A or KITLG was examined by the luciferase reporter gene system. The qRT-PCR detection showed that miR-27a was more highly expressed in white Cashmere goats skin than that in brown Cashmere goats skin. Furthermore, WNT3A and KITLG mRNA and protein expression were down-regulated by miR-27a in vitro and in vivo. A dual-luciferase reporter gene indicated that miR-27a negatively correlates with WNT3A or KITLG. In a word, our research demonstrated that the expression of miR-27a was evidently differential in the white and brown Cashmere goats skin and WNT3A or KITLG was negatively regulated by miR-27a.

Integrated analysis of miRNA and mRNA expression profiles in 2-, 6-, and 12-month-old Small Tail Han Sheep ovaries reveals that oar-miR-432 downregulates RPS6KA1 expression.

Small Tail Han Sheep are an excellent local sheep breed in China, and their outstanding reproductive performance is one of their very important biological characteristics. Clarifying the ovary development process of these ewes should provide a theoretical basis for improving their reproductive efficiency. In this study, we identified the differentially expressed (DE) microRNAs (miRNAs) in 2-, 6-, and 12-month-old small-tail Han sheep ovaries by constructing and analyzing the miRNA expression profiles. These findings clarify the molecular mechanisms regulating the excellent reproductive performance of small-tail Han ewes. We used RNA-Seq technology and bioinformatic to analyze these profiles. Eleven, 13, and 19 DE miRNAs were identified in the 2- vs 6-, 6- vs 12-, and 2- vs 12-month-old ovaries, respectively. In total, 54, 37, and 198 predicted target genes of these DE miRNAs were identified in these three groups, respectively. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analyses showed that in the 2- vs 6-month-old ovaries, the target genes of DE known sheep miRNAs were involved in 102 GO terms and seven signaling pathways; in the 6- vs 12-month-old ovaries, the target genes of DE known sheep miRNAs were involved in 52 GO terms and three signaling pathways; and in the 2- vs 12-month-old ovaries, the target genes of DE known sheep miRNAs were involved in 88 GO terms and six signaling pathways. Three miRNA-target regulatory networks were constructed based on these DE miRNA-target interactions. Nine miRNAs were selected to confirm to the accuracy of the miRNA sequencing data with qRT-PCR. The site at which oar-miR-432 binds RPS6KA1 was determined with a dual-luciferase system. This is the first integrated analysis the expression profiles of miRNAs and their targets during ovarian development in small-tail Han sheep. These data clarify the molecular regulatory mechanisms underlying sheep ovarian development and identify biomarkers that influence the reproductive performance of small-tail Han ewes.

Integrative microRNA-mRNA Analysis of Muscle Tissues in Qianhua Mutton Merino and Small Tail Han Sheep Reveals Key Roles for oar-miR-655-3p and oar-miR-381-5p.

The Qianhua Mutton Merino (QHMM) is a new variety of sheep (Ovis aries) with improved meat performance compared with the traditional Small Tail Han (STH) sheep variety. We recently reported the transcriptome profiling of longissimus muscle tissues between QHMM and STH sheep. In the present study, we aimed to evaluate key micro (mi)RNA-mRNA networks associated with sheep muscle growth and development. We used miRNA sequencing to obtain longissimus muscle miRNA profiles from QHMM and STH sheep. We identified a total of 153 known sheep miRNAs, of which 4 were differentially expressed (DE) between the 2 sheep varieties. We combined these results with mRNA library data to build an miRNA-mRNA network, including 26 target genes of the 4 DE miRNAs. Gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analyses showed that 26 target genes were significantly enriched in 86 biological processes, including muscle organogenesis, myoblast migration, cell proliferation, and adipose tissue development, and in 9 metabolic pathways, including carbohydrate, nucleotide, and amino acid metabolic pathways. oar-miR-655-3p and its target gene ACSM3 and oar-miR-381-5p and its target gene ABAT were selected for subsequent analysis based on GO and KEGG analyses. The binding sites of oar-miR-655-3p with ACSM3 and oar-miR-381-5p with ABAT were validated by a dual-luciferase reporter gene detection system. This represents the first integrative analysis of miRNA-mRNA networks in QHMM and STH muscles and suggests that DE miRNAs, especially oar-miR-655-3p and oar-miR-381-5p, play crucial roles in muscle growth and development.

Differential expression of MC1R gene in Liaoning Cashmere goats with different coat colors.

Melanocyte stimulating hormone receptor (MC1R) has been known as a regulator of eumelanin and phaeomelanin production in the melanocytes, and MC1R mutations causing coat color changes are known in many vertebrates; however, there are no research reports about the differentially expression of MC1R gene and its coding protein in Cashmere goats with different coat color. We examined the presence of MC1R distribution and MC1R protein and gene expression in the white Cashmere goats and black Cashmere goats, respectively; q-PCR, Western blot and immunhistochemical analysis showed that the expression of the MC1R gene in the black Cashmere goats was 3.39 fold more than the white ones (p < 0.01), and Cashmere goats with black genotype had significantly higher (2.03, p < 0.01) MC1R protein expression than white genotype in the all investigated samples. Moreover, all Cashmere goats with different coat color available for immunhistochemical analysis showed either lower (white Cashmere goats) or higher (black Cashmere goats) expression of the MC1R protein; these findings suggested that it had a relationship between the MC1R and the coat color of Cashmere goats. That could lay the foundation for the further research of the MC1R and coat color controllability regulation of the Cashmere goats.

Integrated Analysis of miRNA and mRNA Expression Profiles Reveals Functional miRNA-Targets in Development Testes of Small Tail Han Sheep.

Small Tail Han Sheep is a highly valued local breed in China because of their precocity, perennial estrus, and high fecundity. The average annual lambing rate of ewes is as high as 180-270%, the semen of ram has characteristics of high yield, high density, and good motility. To reveal the key miRNAs and miRNA-targets underlying testis development and spermatogenesis in male Small Tail Han Sheep, integrated analysis of miRNA and mRNA expression profiles in 2-, 6-, and 12-month-old testes was performed by RNA-seq technology and bioinformatics methods. The results showed that total of 153 known sheep miRNAs and 2712 novel miRNAs were obtained in 2-,6 - and 12-month-old Small Tail Han Sheep testes; 5, 1, and 4 differentially expressed (DE) known sheep miRNAs, and 132, 105, and 24 DE novel miRNAs were identified in 2- vs. 6-, 6- vs. 12-, and 2- vs. 12-month-old testes, respectively. We combined miRNA results of this study and the mRNA results obtained in our previous study to predict the target mRNAs of DE known sheep miRNAs; 131, 10, and 15 target mRNAs of DE known sheep miRNAs and 76, 1, and 11 DE miRNA-targets were identified in the three groups, respectively. GO and KEGG analyses showed that: in 2- vs. 6-month-olds, the target genes of DE known sheep miRNAs were involved in 100 biological processes and 11 signaling pathways; in 6- vs. 12-month-olds, the target genes of DE known sheep miRNAs were involved in 4 biological processes; and in 2- vs. 12-month-olds, the target genes of DE known sheep miRNAs were involved in 17 biological processes and 4 signaling pathways. Three miR-target regulatory networks were constructed based on these DE miRNA-targets. The key miRNA-Targets involved in testis development and spermatogenesis were screened. 6 known sheep miRNAs and 6 novel miRNAs were selected to validate the accuracy of miRNA sequencing data by qRT-PCR. The binding sites of oar-miR-379-5p with WNT8A was validated by a dual luciferase reporter gene detection system.

Genetic diversity of the Chinese goat in the littoral zone of the Yangtze River as assessed by microsatellite and mtDNA.

The objective of this study was to assess the genetic diversity and population structure of goats in the Yangtze River region using microsatellite and mtDNA to better understand the current status of those goat genetic diversity and the effects of natural landscape in fashion of domestic animal genetic diversity. The genetic variability of 16 goat populations in the littoral zone of the Yangtze River was estimated using 21 autosomal microsatellites, which revealed high diversity and genetic population clustering with a dispersed geographical distribution. A phylogenetic analysis of the mitochondrial D-loop region (482 bp) was conducted in 494 goats from the Yangtze River region. In total, 117 SNPs were reconstructed, and 173 haplotypes were identified, 94.5% of which belonged to lineages A and B. Lineages C, D, and G had lower frequencies (5.2%), and lineage F haplotypes were undetected. Several high-frequency haplotypes were shared by different ecogeographically distributed populations, and the close phylogenetic relationships among certain low-frequency haplotypes indicated the historical exchange of genetic material among these populations. In particular, the lineage G haplotype suggests that some west Asian goat genetic material may have been transferred to China via Muslim migration.

Differential expression of miR-let7a in hair follicle cycle of Liaoning cashmere goats and identification of its targets.

In order to improve the production and quality of Chinese cashmere, the research of hair follicle development has aroused more and more attention; the regulation mechanism of miRNA in hair follicle development has become a hot spot. A survey of transcriptome profiling screened 10 hair follicle-related miRNAs that were differentially expressed, including miR-let7a. In this study, the expression of miR-let7a was lower in anagen of hair follicle of cashmere goats than that in catagen of hair follicle of cashmere goats (p < 0.01). Results were in accordance with transcriptome data. The expression patterns of miR-let7a target genes (IGF-1R, C-myc, and FGF5) were verified by qRT-PCR, which were consistent with the results of Western blot and showed a downward trend. The dual-luciferase reporter gene system was used to verify the correlation between the expression of miR-let7a and its target genes, and it showed that miR-let7a negatively correlates with C-myc and FGF5. Present study offers new information on miRNAs and their related target genes in the regulation of hair follicle development mechanism.