The neomycin-resistant gene (neo(r)) is probably the most commonly used selectable marker gene in gene targeting and gene transfection research. In this study, the neo(r) gene construct was introduced into in vitro cultured goat foetal fibroblast cells (IV-5), and the cells were selected with 900 microg/ml G418. The G418-resistant colonies were analysed by neo-specific PCR, karyotyping and anti-intermediate filament proteins antibody (anti-vimentin) staining. Cell cycle analysis of the neo(r) positive foetal fibroblast cell colony (IV-5.1) cultured in a variety of cell cycle-arresting medium indicated that 74.2% of cells cultured in serum-deprived medium for 3 days and 71.7% of cells grown to confluence were at G0/G1 stage of cell cycle, respectively, in comparison to 61.6% of cells in normal culture (cycling) medium. Nocodazole treatment for 17 hr in vitro culture could increase the number of cells at G2/M stage of cell cycle from 20.3% (in cycling medium) to 39.7%. In total, one early pregnancy was observed by B ultra-sound scanning in a surrogate transferred with cloned embryos from IV-5.1 cells at M stage (cells were cultured in nocodazole medium). Seven cloned goats, including two that miscarried at a late stage, were derived from the IV-5.1 cell clone cultured in starved medium (G0). Indeed, one surrogate receiving three blastocysts reconstituted from the starved donor cells, gave birth to three live cloned goats, all of which are healthy and doing well. PCR, Southern blot and G418 resistance in vitro of fibroblast cells from cloned goats confirmed that all cloned goats are positive for neo(r) transgene. This study demonstrates that a foreign gene, such as the neo-resistant gene, can be introduced into goat foetal fibroblast cells, and that the resulting transgenic cells are capable of being cloned to produce 100% transgenic animals.

MicroRNA-378 (miRNA-378) has been reported to have a crucial role in skeletal muscle differentiation; however, the underlying mechanisms have largely remained to be elucidated. The present study employed high‑throughput RNA sequencing to investigate the transcriptome following transfection of miRNA‑378 mimics or control RNAs into C2C12 myoblast cells. By sequencing and annotation, 2,802 transcripts that were changed by >1.5 fold were obtained and then subjected to signaling pathway enrichment and gene ontology analysis. Eight genes associated with development were subsequently selected for validation by quantitative qPCR, the results of which were highly consistent with those of the high‑throughput RNA sequencing. The protein levels of bone morphogenetic protein 4 (BMP4), which was among the differentially expressed genes, were decreased following ectopic expression of miRNA‑378. BMP4 was further confirmed to be a direct target of miRNA‑378 by using a dual luciferase assay. Finally, treatment with miRNA‑378 or small interfering RNA against BMP4 induced myogenic differentiation in C2C12 cells. In conclusion, the present study suggested that miRNA‑378 is critical for the promotion of myoblast differentiation by targeting BMP4.

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Multidrug resistance (MDR) is an increasing public health concern worldwide. Artesunate (ART) has been reported to be significantly effective in enhancing the effectiveness of various b-lactam antibiotics against MDR Escherichia coli via inhibiting the efflux pump genes. Apart from b-lactam antibiotics, there is no report regarding the potential synergistic effects of ART combining with fluoroquinolones (FQs). In this study, we investigated whether ART can enhance the antibacterial effects of FQs in vitro. The antibacterial activity of ART and antibiotics against 13 animal-derived E. coli clinical isolates was assessed for screening MDR strains. Then the synergistic activity of FQs with ART against MDR E. coli isolates was evaluated. Daunorubicin (DNR) accumulation within E. coli and messenger RNA (mRNA) expressions of acrA, acrB, tolC, and qnr genes were investigated. The results showed that ART did not show significant antimicrobial activity. However, a dramatically synergistic activity of ART combining with FQs was obsessed with (SFIC) = 0.12-0.33. ART increased the DNR accumulation and reduced acrAB-tolC mRNA expression, but enhanced the mRNA expression of qnrS and qnrB within MDR E. coli isolates. These findings suggest that ART can potentiate FQs activity which may be associated with drug accumulation by inhibiting the expression of acrAB-tolC.

Skeletal muscle accounts for ~40% of total body mass. The principle functions of skeletal muscle include supporting the body structure, controlling motor movements and storing energy. Rhabdomyosarcoma (RMS) is a skeletal muscle‑derived soft tissue tumor widely occurring in the pediatric population. In previous years, microRNAs (miRNAs) have been demonstrated to be important in skeletal muscle development, function and the pathogenesis of various diseases, including RMS. The present review provided an overview of current knowledge on the muscle‑specific and ubiquitously‑expressed miRNAs involved in skeletal muscle differentiation and their dysregulation in RMS. Additionally, the potential use and challenges of miRNAs as therapeutic targets in this soft‑tissue sarcoma were examined and the future prospects for miRNAs in muscle biology and muscle disorders were discussed.