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目的获得具备从内含体逃逸的宫颈癌特异性穿膜肽GFP-HA-PTP融合蛋白,探讨其在宫颈癌细胞中的靶向穿膜效应。方法构建p ET15b-GFP-HA-PTP融合原核表达质粒,并转化大肠杆菌BL21,经IPTG诱导和NI-NTA树脂纯化得到融合蛋白,经Western blotting分析确认后,于荧光显微镜下观察GFP-HA-PTP的靶向穿膜活性。结果大肠杆菌高效表达了经Western blot鉴定正确的FP-HA-PTP融合蛋白,且此融合蛋白能够靶向宫颈癌细胞株Siha和Caski,并高效地从内含体中逃逸进入胞浆。结论 GFP-HA-PTP融合蛋白能够高效靶向穿膜进入培养宫颈癌细胞。
Objective To obtain GFP-HA-PTP fusion protein, which escapes from inclusion bodies, and to investigate its targeted transmembrane effect in cervical cancer cells. METHODS: The fusion protein p ET15b-GFP-HA-PTP fusion prokaryotic expression vector was constructed and transformed into E.coli BL21. The fusion protein was induced by IPTG induction and NI-NTA resin purification. The fusion protein was confirmed by Western blotting. Targeting transmembrane activity of PTP. Results Escherichia coli efficiently expressed the FP-HA-PTP fusion protein identified by Western blot, and the fusion protein could target cervical cancer cell lines Siha and Caski and efficiently escape from the inclusion body into the cytoplasm. Conclusion The GFP-HA-PTP fusion protein can efficiently target the transmembrane into the culture of cervical cancer cells.