Principal component analysis-artificial neural network-based model for predicting the static strength of seasonally frozen soils.

Seasonally frozen soils are exposed to freeze‒thaw cycles every year, leading to mechanical property deterioration. To reasonably describe the deterioration of soil under different conditions, machine learning (ML) technology is used to establish a prediction model for soil static strength. Six key influencing factors (moisture content, compaction degree, confining pressure, freezing temperature, number of freeze‒thaw cycles and thawing duration) are included in the modelling database. The accuracy of three typical ML algorithms (support vector machine (SVM), random forest (RF) and artificial neural network (ANN)) is compared. The results show that the ANN outperforms the SVM and RF. Principal component analysis (PCA) is combined with the ANN, and the PCA-ANN algorithm is proposed, which further improves the prediction accuracy. The deterioration of soil static strength is systematically researched using the PCA-ANN algorithm. The results show that the soil static strength decreased considerably after the first several freeze‒thaw cycles before the strength plateau occurred, and the strength reduction increased significantly with increasing moisture content and compaction degree. The PCA-ANN model can generate a reasonable prediction for the static strength or other soil properties of seasonally frozen soil, which will provide a scientific reference for practical engineering.

Study on the SHP2-Mediated Mechanism of Promoting Spermatogenesis Induced by Active Compounds of Eucommiae Folium in Mice.

Spermatogenesis directly determines the reproductive capacity of male animals. With the development of society, the increasing pressure on people's lives and changes in the living environment, male fertility is declining. The leaf of Eucommia ulmoides Oliv. (Eucommiae Folium, EF) was recorded in the 2020 Chinese Pharmacopoeia and was used in traditional Chinese medicine as a tonic. In recent years, EF has been reported to improve spermatogenesis, but the mechanisms of EF remain was poorly characterized. In this study, the effect of EF ethanol extract (EFEE) on spermatogenesis was tested in mice. Chemical components related to spermatogenesis in EF were predicted by network pharmacology. The biological activity of the predicted chemical components was measured by the proliferation of C18-4 spermatogonial stem cells (SSCs) and the testosterone secretion of TM3 leydig cells. The biological activity of chlorogenic acid (CGA), the active compound in EF, was tested in vivo. The cell cycle was analysed by flow cytometry. Testosterone secretion was detected by ELISA. RNA interference (RNAi) was used to detect the effect of key genes on cell biological activity. Western blotting, qRT-PCR and immunofluorescence staining were used to analyse the molecular mechanism of related biological activities. The results showed that EFEE and CGA could improve spermatogenesis in mice. Furthermore, the main mechanism was that CGA promoted SSC proliferation, self-renewal and Leydig cell testosterone secretion by promoting the expression of SHP2 and activating the downstream signaling pathways involved in these biological processes. This study provided strong evidence for elucidating the mechanism by which EF promotes the spermatogenesis in mice and a new theoretical basis for dealing with the decrease in male reproductive capacity.

Coronavirus GenBrowser for monitoring the transmission and evolution of SARS-CoV-2.

Genomic epidemiology is important to study the COVID-19 pandemic, and more than two million severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) genomic sequences were deposited into public databases. However, the exponential increase of sequences invokes unprecedented bioinformatic challenges. Here, we present the Coronavirus GenBrowser (CGB) based on a highly efficient analysis framework and a node-picking rendering strategy. In total, 1,002,739 high-quality genomic sequences with the transmission-related metadata were analyzed and visualized. The size of the core data file is only 12.20 MB, highly efficient for clean data sharing. Quick visualization modules and rich interactive operations are provided to explore the annotated SARS-CoV-2 evolutionary tree. CGB binary nomenclature is proposed to name each internal lineage. The pre-analyzed data can be filtered out according to the user-defined criteria to explore the transmission of SARS-CoV-2. Different evolutionary analyses can also be easily performed, such as the detection of accelerated evolution and ongoing positive selection. Moreover, the 75 genomic spots conserved in SARS-CoV-2 but non-conserved in other coronaviruses were identified, which may indicate the functional elements specifically important for SARS-CoV-2. The CGB was written in Java and JavaScript. It not only enables users who have no programming skills to analyze millions of genomic sequences, but also offers a panoramic vision of the transmission and evolution of SARS-CoV-2.

Genetic Features of Plasmid- and Chromosome-Mediated mcr-1 in Escherichia coli Isolates From Animal Organs With Lesions.

The genomic context of the mcr-1 gene in Escherichia coli from animal feces has been widely reported. However, less is known about the mcr-1-carrying plasmid characteristics and other functional regions of Escherichia coli isolates from animal organs with lesions. The present study investigated the antimicrobial resistance, population structure, and genetic features of mcr-1-positive Escherichia coli strains isolated from animal organs with lesions. The antimicrobial susceptibility testing indicated that 24 mcr-1-positive Escherichia coli isolates were resistant to at least three or all antimicrobial categories. MLST analysis suggested that the dominant clone complexes (CC) were mainly CC156, CC448, and CC10. In addition, ST10596, a newly discovered sequence type in swine, failed to be classified. Meanwhile, the mcr-1 gene located on the different plasmids was successfully transferred to the recipients, and whole-genome sequencing indicated the mcr-1 gene was embedded in mcr-1-pap2 cassette but not flanked by ISApl1. The mcr-1 gene is located on the chromosome and embedded in Tn6330. Furthermore, NDM-5 was found on the IncX3-type plasmid of J-8. The virB6 and traI gene of type IV secretion system (T4SS) were truncated by IS2 and IS100 and located on the IncX4- and the IncHI2/HI2A/N-type plasmids, respectively. The multidrug-resistant (MDR) region of IncHI2/HI2A/N-type plasmids contained two class 1 integrons (In0, In640) and four composite transposons (Tn4352, Tn6010, cn_4692_IS26, cn_6354_IS26). Overall, 24 mcr-1-positive Escherichia coli isolates in our study showed MDR, or even extensively drug resistant (XDR), and exhibited population diversity. The T4SS gene truncation by the insertion sequence may affect the efficiency of plasmid conjugative transfer. Furthermore, the class 1 integrons and composite transposons in the MDR region of IncHI2/HI2A/n-type plasmid contributed to the multireplicon plasmid formation, the acquisition, and transfer of antimicrobial resistance genes (ARGs).

Bioactivity-guided isolation of immunomodulatory compounds from the fruits of Ligustrum lucidum.

ETHNOPHARMACOLOGICAL RELEVANCE: The fruits of Ligustrum lucidum (FLL) W.T. Aiton (Oleaceae) is included in the 2020 "Chinese Pharmacopoeia" and is widely used in traditional Chinese medicine as a tonic. In recent years, FLL has been reported to improve immune function, but the bioactive compounds and mechanisms of FLL remain poorly characterized. AIM OF THE STUDY: To identify FFL compounds with strong immune activity and explore their molecular mechanisms. MATERIALS AND METHODS: The phagocytic activity of RAW264.7 macrophages and proliferation activity of spleen lymphocytes were used to guide the isolation of bioactive compounds from FLL extracts. Lymphocyte subpopulations, Ca2+ concentrations, and surface molecule expression were analyzed using flow cytometry. Cytokine secretion was examined using ELISA. FITC-OVA uptake was observed using fluorescence microscopy. NF-κB activation was analyzed using western blotting. RESULTS: The extraction and isolation produced ten compounds, namely oleuropeinic acid, nuezhenide, isonuezhenide, salidroside, isoligustrosidic acid, ligulucidumosides A, 8(E)-nuezhenide, hydroxytyrosol, oleuropein, and p-hydroxyphenethyl 7-ß-D-glucosideelenolic acid ester were isolated and identified from FLL-Bu-30%. Immunoactivity experiments showed that hydroxytyrosol had the strongest macrophage phagocytotic and lymphocyte proliferation-promoting activities. Further studies showed that hydroxytyrosol could significantly enhance lymphocyte subsets CD3+, CD4+/CD8+, and CD3+CD4-CD8-, promote IL-4, IFN-γ, and TNF-α secretion, and increase intracellular Ca2+ concentrations. In addition, the results from RAW264.7 macrophages showed that hydroxytyrosol increased FITC-OVA uptake, induced TNF-α and IL-1ß production, upregulated MHC-II, CD80, and CD86 expression, promoted cytoplasmic IκB-α degradation, and increased nuclear NF-κB p65 levels. CONCLUSION: Our study provides substantial evidence regarding the mechanism of the immunomodulatory effects of compounds from FLL.

Melatonin Ameliorates Busulfan-Induced Spermatogonial Stem Cell Oxidative Apoptosis in Mouse Testes.

AIMS: Many men endure immunosuppressive or anticancer treatments that contain alkylating agents before the age of sexual maturity, especially the increasing number of preadolescent males who undergo busulfan treatment for myeloablative conditioning before hematopoietic stem cell transplantation. Before sperm production, there are no sperm available for cryopreservation. Thus, it is necessary to identify a solution to ameliorate the busulfan-induced damage of spermatogonial stem cells (SSCs). RESULTS: In this study, we demonstrated that melatonin relieved the previously described SSC loss and apoptosis in mouse testes. Melatonin increased the expression of manganese superoxide dismutase (MnSOD), which regulated the production of busulfan-induced reactive oxygen species (ROS). Moreover, melatonin promoted sirtuin type 1 (SIRT1) expression. SIRT1 participated in the deacetylation of p53, which promotes p53 ubiquitin degradation. Decreased concentrations of deacetylated p53 resulted in spermatogonial cell resistance to apoptosis. Acute T cell leukemia cell assay demonstrated that melatonin does not affect busulfan-induced cancer cell apoptosis and ROS. INNOVATION: The current evidence suggests that melatonin may alleviate the side effects of alkylating drugs, such as busulfan. CONCLUSION: Melatonin promoted MnSOD and SIRT1 expression, which successfully ameliorated busulfan-induced SSC apoptosis caused by high concentrations of ROS and p53. Antioxid. Redox Signal. 28, 385-400.

p38 MAPK pathway is essential for self-renewal of mouse male germline stem cells (mGSCs).

OBJECTIVES: Male germline stem cells (mGSCs), also called spermatogonial stem cells (SSCs), constantly generate spermatozoa in male animals. A number of preliminary studies on mechanisms of mGSC self-renewal have previously been conducted, revealing that several factors are involved in this regulated process. The p38 MAPK pathway is widely conserved in multiple cell types in vivo, and plays an important role in cell proliferation, differentiation, inflammation and apoptosis. However, its role in self-renewal of mGSCs has not hitherto been determined. MATERIALS AND METHODS: Here, the mouse mGSCs were cultured and their identity was verified by semi-RT-PCR, alkaline phosphatase (AP) staining and immunofluorescence staining. Then, the p38 MAPK pathway was blocked by p38 MAPK-specific inhibitor SB202190. mGSC self-renewal ability was then analysed by observation of morphology, cell number, cell growth analysis, TUNEL incorporation assay and cell cycle analysis. RESULTS: Results showed that mouse mGSC self-renewal ability was significantly inhibited by SB202190. CONCLUSIONS: This study showed for the first time that the p38 MAPK pathway plays a key role in maintaining self-renewal capacity of mouse mGSCs, which offers a new self-renewal pathway for these cells and contributes to overall knowledge of the mechanisms of mGSC self-renewal.

CD49f promotes proliferation of male dairy goat germline stem cells.

OBJECTIVES: CD49f enhances multipotency and maintains stemness in embryonic stem cells (ESCs), however, whether it would be effective in mGSCs has remained unclear. Moreover, better standards for mGSC enrichment and purification are necessary. The present study was conducted to determine roles of CD49f in mGSC enrichment and regulation. MATERIALS AND METHODS: CD49f expression patterns were investigated in dairy goats. CD49f positive cells were purified and enriched using magnetic-activated cell sorting (MACS), and characteristics of the cultured cells were assayed using alkaline phosphatase (AP) analysis, quantitative real-time PCR (QRT-PCR) and immunofluorescence analysis. Furthermore, the exogenous CD49f gene was transfected into mGSCs and its effects were analysed. RESULTS: CD49f was found to be conserved in both mRNA and amino acid sequences and that it was an efficient marker for dairy goat mGSC identification, enrichment and purification. CD49f positive cells expressed higher levels of mGSC-specific markers, and proliferated faster than CD49f negative cells. Overexpression CD49f promoted proliferation of dairy goat mGSCs, and Oct4 expression was upregulated; histone H3-lysine 9 dimethylation (H3K9me2) was reduced. CONCLUSIONS: Taken together, our data suggest that CD49f plays novel and dynamic roles in regulating maintenance of pluripotency in mGSCs via Oct4 crosstalk and histone methylation dynamics,which may provide new solutions for mGSCs stability in vitro.

EIF2S3Y suppresses the pluripotency state and promotes the proliferation of mouse embryonic stem cells.

Eukaryotic translation initiation factor 2, subunit 3, and structural gene Y-linked (EIF2S3Y) is essential for spermatogenesis in mouse models. However, its effect on embryonic stem (ES) cells remains unknown. In our observation, differentiated ES cells showed higher levels of EIF2S3Y. To further elucidate its role in ES cells, we utilized ES-derived EIF2S3Y-overexpressing cells and found that EIF2S3Y down-regulated the pluripotency state of ES cells, which might be explained by decreased histone methylation levels because of reduced levels of ten-eleven translocation 1 (TET1). Moreover, EIF2S3Y-overexpressing cells showed an enhanced proliferation rate, which might be due to increased Cyclin A and Cyclin E levels. This study highlighted novel roles of EIF2S3Y in the pluripotency maintenance and proliferation control of ES cells, which would provide an efficient model to study germ cell generation as well as cancer development using ES cells, thus providing valuable target for clinical applications of ES cells.

PLZF-Induced Upregulation of CXCR4 Promotes Dairy Goat Male Germline Stem Cell Proliferation by Targeting Mir146a.

Previous studies have shown that promyelocytic leukemia zinc finger (PLZF), chemokine (C-X-C motif) receptor 4 (CXCR4) and mir146a were associated with the self-renewal of mouse spermatogonial stem cells (SSCs); however, there is little information on their effects on the fate of livestock SSCs. Here, we have identified a regulatory pathway in dairy goat mGSCs, involving PLZF, mir146a and the SDF-1 receptor CXCR4. PLZF overexpression downregulated mir146a and simultaneously upregulated the expression of CXCR4 protein, whereas PLZF knockdown (siPLZF) induced the specifically opposite effects. The in vitro assays demonstrated that PLZF specifically interacts with and suppresses the mir146a promoter, and mir146a targets CXCR4 to impede its translation. The levels of ERK1/2 phosphorylation in the mGSCs overexpressed CXCR4 and PLZF were upregulated, respectively, whereas mir146a expression was decreased and CXCR4 protein was increased. Mir146a overexpression and siPLZF impaired mGSC proliferation and differentiation, however, Mir146a knockdown induced the opposite effects. The effects of PLZF and mir146a were mediated regulation by mir146a and CXCR4, respectively. Overexpression of CXCR4 or addition of CXCL12 in cultures of dairy goat mGSCs resulted in the upregulation of their signaling, and the phosphorylation of ERK1/2 was increased. Collectively, these findings indicate that PLZF is an important transcription factor in the regulation of the expression of CXCR4 to promote dairy goat mGSC proliferation by targeting mir146a.

miR-204 Regulates the Proliferation of Dairy Goat Spermatogonial Stem Cells via Targeting to Sirt1.

The regulation of spermatogonial stem cell (SSC) proliferation and self-renewal is a complex process. Several studies on the microRNA regulation of mammalian spermatogenesis have been reported. Here, we predicted miRNA targeting of Sirt1, and a dual luciferase experiment confirmed that miR-204 interacted with the Sirt1 3'-untranslated region (3'-UTR). The expression of miR-204 and Sirt1 in dairy goat testicles was investigated, and the results showed that the expression pattern of Sirt1 was similar to that of miR-204 in the temporal-spatial distribution. The over-expression of Sirt1 in goat SSCs can promote SSCs' self-renewal gene expression and cell proliferation. Furthermore, miRNA sequencing results showed that Sirt1 had a higher expression level in dairy goat CD49f(+) and CD90(+) SSCs, but the expression level of miR-204 was lower. In an in vitro assay, Sirt1 was significantly down-regulated in dairy goat SSCs when transfected with miR-204 mimics, indicating that Sirt1 was a target of miR-204 in the dairy goat. On the basis of the results of RT-qPCR, fluorescence-activated cell sorting (FACS), and western blotting, we found that the over-expression of Sirt1 in goat SSCs can promote cellular proliferation and change self-renewal and pluripotent gene expression. Thus, miR-204 was involved in the regulation of dairy goat SSCs proliferation via Sirt1.

Ras/ERK1/2 pathway regulates the self-renewal of dairy goat spermatogonia stem cells.

Spermatogonia stem cells (SSCs), also named the male germline stem cells (mGSCs), which is located at the base of the seminiferous tubules of testis, is the basis for generating sperm steadily in male animals. Currently, there are some preliminary study on the self-renewal and differentiation of SSCs, but further mechanism, especially in large animals, has not been clearly understood. Ras/ERK1/2 pathway is widely distributed in multiple cells in vivo. It plays an important role in cell proliferation, differentiation and so on. However, the study on the function for the self-renewal of dairy goats SSCs has not been investigated. In this study, the dairy goat SSCs characterization were evaluated by semi-RT-PCR, alkaline phosphatase (AP) staining, and immunofluorescence staining. Then, Ras/ERK1/2 pathway was blocked by specific MEK1/2 inhibitor PD0325901. We analyzed the proliferation by cell number, cell growth curve, Brdu incorporation assay, and cell cycle analysis. The results showed that the proliferation was significantly inhibited by PD0325901. Cell apoptosis induced by blocking the Ras/ERK1/2 pathway was analyzed by TUNEL. The expression of ETV5 and BCL6B, the downstream gene of Ras/ERK1/2 pathway, was downregulated. This study suggest that the Ras/ERK1/2 pathway plays a critical role in maintaining the self-renewal of dairy goat SSCs via regulation of ETV5 and BCL6B. This study laid a foundation for insights into the mechanism of SSCs self-renewal comprehensively.

miR-544 Regulates Dairy Goat Male Germline Stem Cell Self-Renewal via Targeting PLZF.

The balance between the self-renewal and differentiation of male germline stem cells (mGSCs) is critical for the initiation and maintenance of mammalian spermatogenesis. The promyelocytic leukemia zinc finger (PLZF), a zinc finger protein, is a critical factor for maintaining the self-renewal of mGSCs, so, evaluation of the PLZF pathway in mGSCs may provide a deeper insight into mammalian spermatogenesis. miRNA was also an important regulating factor for the self-renewal and differentiation of mGSCs; however, there is currently no data indicating that which miRNA regulate the self-renewal and differentiation of mGSCs via PLZF. Here, we predicted the prospective miRNA targeting to PLZF using the online Bioinformatics database-Targetscan, and performed an analysis of the dual-luciferase recombinant vector, psiCHCEKTM-2-PLZF-3'UTR. miR-544 mimics (miR-544m), miR-544 inhibitors (miR-544i), Control (NC, scrambled oligonucleotides transfection), pPLZF-IRES2-EGFP or PLZF siRNA were transfected into mGSCs; the cells proliferation was evaluated by BRDU incorporation assay and flow cytometry, and the mGSC marker, GFRa1, PLZF, KIT, DAZL, and VASA expression were analyzed by RT-qPCR, immunofluorescence and Western blot. The results showed that miR-544 regulates dairy goat male germline stem cell self-renewal via targeting PLZF. Our study identifies a new regulatory pathway for PLZF and expands upon the PLZF regulatory network in mGSCs.

The Tet1 and histone methylation expression pattern in dairy goat testis.

DNA methylation and histone methylation are critical for mammalian development. Ten-eleven translocation (Tet1), a key regulator of DNA methylation, has been identified as a key enzyme for the activation of DNA demethylation; histone H3 lysine 9 (H3K9) and 27 (H3K27) methylation repress gene expression. Significant progress on the biological functions of Tet proteins has been made in mice and humans. However, their expression pattern and function in the male germ cells in the dairy goat testis are still unclear. The present study described the expression pattern of Tet1, H3K9, and H3K27 in the dairy goat testis and cultured goat spermatogonia stem cells (gSSCs). The results showed that Tet1 was weakly expressed in the dairy goat's testis compared to other organ tissues. Tet1, 5-hydroxymethylcytosine, H3K9, and H3K27 expressions were positive and dynamically changing during spermatogenesis; however, they showed weak expression in neonate stage in vivo. Tet1 and 5-hydroxymethylcytosine showed low expression in gSSCs in vitro in differentiated cultures. These will provide new perspectives for DNA methylation/demethylation and better regulation of epigenetic modifications in gSSCs.

CD49f-positive testicular cells in Saanen dairy goat were identified as spermatogonia-like cells by miRNA profiling analysis.

miRNAs, a type of small RNA, play critical roles in mammalian spermatogenesis. Spermatogonia are the foundation of spermatogenesis and are valuable for the study of spermatogenesis. However, the expression profiling of the miRNAs in spermatogonia of dairy goats remains unclear. CD49f has been one of the surface markers used for spermatogonia enrichment by magnetic activated cell sorting (MACS). Therefore, we used a CD49f microbead antibody to purify CD49f-positive and -negative cells of dairy goat testicular cells by MACS and then analysed the miRNA expression in these cells in depth using Illumina sequencing technology. The results of miRNA expression profiling in purified CD49f-positive and -negative testicular cells showed that 933 miRNAs were upregulated in CD49f-positive cells and 916 miRNAs were upregulated in CD49f-negative cells with a twofold increase, respectively; several miRNAs and marker genes specific for spermatogonial stem cells (SSCs) in testis had a higher expression level in CD49f-positive testicular cells, including miR-221, miR-23a, miR-29b, miR-24, miR-29a, miR-199b, miR-199a, miR-27a, and miR-21 and CD90, Gfra1, and Plzf. The bioinformatics analysis of differently expressed miRNAs indicated that the target genes of these miRNAs in CD49f-positive cells were involved in cell-cycle biological processes and the cell-cycle KEGG pathway. In conclusion, our comparative miRNAome data provide useful miRNA profiling data of dairy goat spermatogonia cells and suggest that CD49f could be used to enrich dairy goat spermatogonia-like cells, including SSCs.

miR-34c enhances mouse spermatogonial stem cells differentiation by targeting Nanos2.

miRNAs are expressed in many mammalian cells, acting specific roles in regulating gene expression or mediating special mRNAs cleavage by targeting their 3'-untranslated region (3'UTR). Some miRNAs are essential and important for animal development. However, it is still unclear what the relationship is between miR-34c and mammalian spermatogonial stem cells (SSCs). We found that a conserved microRNA-34c through its target-Nanos2, regulating SSCs' differentiation in mouse. Immunohistochemistry analysis of Nanos2 and miR-34c FISH results revealed the opposite expression trends between them. Seven bioinformatics websites and programs predicted that miR-34c has interaction sites in Nanos2's 3'UTR. Dual-luciferase reporter vector and mutated dual-luciferase reporter vector analysis validated that they are interacted. After transfection miR-34c mimics into mouse SSCs, or miR-34c lentiviral vector in vitro co-cultivation with seminiferous tubules, and Western blot analysis demonstrated that miR-34c over-expression could suppress Nanos2 expression in post-transcription level. Our experiments identified that miR-34c may promote meiosis process by interacting with Nanos2 leading up-regulation of Stra8 in mouse spermatogonial stem cells.

The function of Msx1 gene in promoting meiosis of dairy goat male germline stem cells (mGSCs).

During sequential stages of meiosis, numerous cytoplasmic and nuclear events take place in which many germline and non-germline genes involved. It is demonstrated that the germline gene Stra8 and synaptonemal complex protein 3 (Scp3) play an important role in the meiosis. Recently, studies showed Msx1, a DNA-binding protein taking part in the skeletal development, also having a functional attractive factor to Stra8 and Scp3 in the meiosis. In this study, we cloned the gene Msx1 then transfected the Msx1 constructed recombination plasmid, pMsx1-Ires2-AcGFP, into the dairy goat germline stem cells (male germline stem cells) and analysed the effects of Msx1 on the expression of Stra8 and Scp3. The results showed that Msx1 could enhance the expression of Stra8 and Scp3 and promote the meiosis in goat testicular cells. Bmp4 activated the expression of Msx1 and Stra8. This study suggests that Msx1 plays an important role in spermatogenesis and meiosis.

Enrichment and characterization of Thy1-positive male germline stem cells (mGSCs) from dairy goat (Capra hircus) testis using magnetic microbeads.

In mammalian testis, male germline stem cells (mGSCs) are originated from primordial germ cells and developed into spermatocyte and spermatid. In our previous studies, we had isolated a pluripotent mGSCs from goat testes and tested their pluripotency and differentiation potential in vitro and in vivo, which revealed that the isolated and cultured dairy goat mGSCs maintained the characteristics of mGSCs. However, Thy1, a marker of mGSCs, was not examined in detail. In this study, the dairy goat mGSCs were purified by differential plating followed by magnetic-activated cell sorting (MACS) using Thy1 antibody. The quantitative reverse transcription polymerase chain reaction and immunofluorescence analyses revealed that the transcription and expression of Thy1, CD49f, Plzf, Oct4, Gfra1, and Vasa were higher in Thy1-positive cells when compared with Thy1-negative cells. The detection results of culturing dairy goat mGSCs indicated that the Thy1-positive cells maintained the characteristics of mGSCs, grew relatively faster than Thy1-negative cells, and the percentage of alkaline phosphatase-positive cells and colonies were significantly higher in Thy1-positive mGSCs than Thy1-negative cells. Collectively, these results indicate that THY1 is a marker of undifferentiated spermatogonia in goat testes, the technique of magnetic-activated cell sorting using Thy1 antibody could be an efficient method to enrich mGSCs in goat.