Bin Liu, Shichao Wang, Fanhua Meng, Bei Wu, Yanru Zhang, Junwei Cao
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引用次数: 0
Abstract
This study aims to establish an immortalized fibroblast cell line from Mongolian sheep. Primary Mongolian sheep fibroblasts (SSF) were isolated using tissue explant and enzymatic digestion methods, followed by microscopic observation, growth curve plotting, and karyotype analysis. The results confirmed the successful isolation of SSF. Human (hTERT) and sheep (sTERT) telomerase reverse transcriptase vectors were separately introduced into SSF, with cells passaged up to 36 generations following G418 selection. Microscopic examination and qRT-PCR results demonstrated that TERT transfection did not alter the morphology of SSF and led to stable, high levels of TERT expression (P < 0.01). Cell counting and flow cytometry revealed that TERT-transfected cells had higher viability and lower apoptosis rates compared to SSF (P < 0.05). Karyotype and soft agar colony formation assays indicated that hTERT and sTERT-transfected cells maintained normal characteristics without malignant transformation. β-galactosidase staining indicated that TERT transfection significantly reduced cellular senescence (P < 0.001). Additionally, sTERT-transfected cells exhibited higher TERT expression, enhanced viability, proliferation, and anti-senescence effects compared to hTERT-transfected cells (P < 0.05). In summary, the introduction of hTERT and sTERT effectively extends the lifespan of SSF, with sTERT demonstrating a more pronounced effect. This study provides critical evidence for preserving Mongolian sheep genetic resources and developing immortalized cell lines.
本研究旨在建立蒙古羊成纤维细胞永生化系。采用组织外植体法和酶消化法分离蒙古羊原代成纤维细胞(SSF),进行显微镜观察、生长曲线绘制和核型分析。结果证实SSF的成功分离。将人(hTERT)和羊(sTERT)端粒酶逆转录酶载体分别导入SSF,在G418选择后,细胞传代达36代。显微镜检查和qRT-PCR结果表明,TERT转染没有改变SSF的形态,并导致稳定、高水平的TERT表达(P P P P
期刊介绍:
Biotechnology can be defined as any technique that uses living organisms (or parts of organisms like cells, genes, proteins) to make or modify products, to improve plants, animals or microorganisms for a specific use. Animal Biotechnology publishes research on the identification and manipulation of genes and their products, stressing applications in domesticated animals. The journal publishes full-length articles and short research communications, as well as comprehensive reviews. The journal also provides a forum for regulatory or scientific issues related to cell and molecular biology applied to animal biotechnology.
Submissions on the following topics are particularly welcome:
- Applied microbiology, immunogenetics and antibiotic resistance
- Genome engineering and animal models
- Comparative genomics
- Gene editing and CRISPRs
- Reproductive biotechnologies
- Synthetic biology and design of new genomes