猪骨骼肌肌内脂肪（IMF）含量是肉质性状重要评定指标之一，肌内脂肪细胞来源于中胚层间充质干细胞特化的纤维/脂肪形成祖细胞（FAPs），FAPs细胞的增殖能力与IMF含量正相关。TG结合因子1（TGIF1）是转录辅阻遏物，为研究其对猪FAPs细胞增殖能力的调控，本项目首先拟利用单细胞转录组测序技术鉴定猪背最长肌FAPs细胞特异表达表面抗原基因，然后利用流式细胞分选技术分离猪FAPs细胞，并评价其体外传代能力，通过培养基的优化和小分子药物筛选获得体外可稳定传代的猪FAPs细胞。再利用CRISPR/Cas9技术构建条件性超表达和敲除TGIF1基因的细胞系，结合ChIP-seq和RNA-seq技术，识别TGIF1直接抑制的与细胞增殖相关的基因，并通过蛋白质谱技术鉴别TGIF1互作的转录抑制因子及其对细胞增殖相关基因的调控，以期阐明TGIF1调控猪FAPs细胞增殖能力的分子机理。

The intramuscular fat (IMF) content of porcine skeletal muscle is one of the important indicators of meat quality traits. Studies have shown that intramuscular adipocytes are derived from mesodermal mesenchymal stem cells specialized fibro/adipogenic progenitors (FAPs). The proliferation ability of FAPs cells is positively correlated with IMF content. TG-interacting factor 1 (TGIF1) is a transcriptional corepressor. To study the regulation of TGIF1 on the proliferation ability of porcine FAPs cells, this project first intends to identify the specifically expressed surface antigen genes of porcine FAPs cells within longissimus dorsi muscle using single-cell transcriptome sequencing technology, and the porcine FAPs cells will be isolated via flow cytometry. The in vitro passage ability of porcine FAPs cells will be evaluated and the in vitro stably passaged FAPs cell line will be established by optimizing of medium and screening a batch of small molecule drugs. Conditional overexpression and knockout TGIF1 gene cell lines will be constructed using CRISPR/Cas9 technology. Additionally, the cell proliferation related genes which are directly suppressed by TGIF1 will be identified using ChIP-seq and RNA-seq techniques. Mass-spectrometric technique will be used to discern the TGIF1 interacted transcription repressors. One transcription repressor will be elected to study its effect on the transcriptional regulation of cell proliferation related genes. Ultimately, we are expecting to elucidate the potential molecular mechanism of TGIF1 regulating the proliferation ability of porcine FAPs cells.