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人源Cdc25C蛋白的融合表达及纯化 被引量:2

Prokaryotic Expression and Purification of Human Cdc25C
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摘要 目的:表达并纯化有活性的GST-Cdc25C融合蛋白,以用于Cdc25C功能研究。方法:利用RT-PCR克隆MCF-7细胞的cdc25c全长基因;在大肠杆菌中表达GST-Cdc25C融合蛋白;利用GSH交联的琼脂糖珠纯化GST-Cdc25C融合蛋白;通过体外磷酸酶活性分析检测GST-Cdc25C融合蛋白的磷酸酶活性。结果:克隆获得1465 bp的人源cdc25c全长基因,并克隆至pGEX-4T-1原核表达载体;在原核系统中可溶性表达了相对分子质量约87×103的GST-Cdc25C融合蛋白;通过亲和纯化获得的GST-Cdc25C融合蛋白具有较好的磷酸酶活性。结论:得到了有磷酸酶活性的GST-Cdc25C融合蛋白,可用于后续的Cdc25C功能研究。 Objective: To prokaryotic express and purify GST-Cdc25C fusion proteins with bioactivity. Methods: The full-length of human cdc25c gene was cloned from MCF-7 ceils by RT-PCR, then it was inserted into plas- mid pGEX-4T-1 and expressed in E.coli. The GST-Cdc25C fusion protein was purified by method of GSH-aga- rose beads affinity, and its phosphatase activity was measured by using 3-OMFP substrates in vitro. Results: cdc25c gene with 1465 bp was cloned into pGEX-4T-1 vector, and GST-Cdc25C fusion protein about 87 kD was expressed and purified. The GST-Cdc25C fusion protein showed remarkable phosphatase activity. Conclusion: GST-Cdc25C fusion protein with bioactivity was acquired, and it could be used for Cdc25C function research.
出处 《生物技术通讯》 CAS 2013年第4期481-483,共3页 Letters in Biotechnology
关键词 Cdc25C 融合表达 磷酸酶活性 Cdc25C fusion expression phosphatase activity
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同被引文献39

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