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| Expression Vector Construction of Staphylococcus aureus Histidine Kinase AgrC Based on RF Cloning Combined with Nested PCR |
| LU Lu1,2, XIONG Wen1,2, ZHANG Yu-kun1,2, WANG Li-na3, QUAN Chun-shan1,2, FAN Sheng-di1,2 |
1. SEAC-ME Key Laboratory, Dalian 116600, China;
2. Department of Life Science, Dalian Nationalities University, Dalian 116600, China;
3. Dalian Institute of Chemical Physics, Chinese Academy of Sciences, Dalian 116023, China |
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Abstract AgrC is a membrane-embedded histidine kinase of Staphylococcus aureus that is thought to act as a sensor for the recognition of environmental signals and the transduction of signals into the cytoplasm so as to regulate and control a series of related pathogenic gene expression. Restriction-Free cloning and Nested PCR were used to construct an expression vector of pET-28a-AgrC successfully. Then, expression vector pET-28a-AgrC was transformed into E.coli C43 (DE3), and then, IPTG was added to induce expression. The expressed AgrC protein with a C-terminal His-tagged was purified using immobilized metal affinity chromatography (IMAC) and size exclusion chromatography (SEC). SDS-PAGE test showed that AgrC proteins were successfully separated and purified.
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Received: 25 June 2014
Published: 25 October 2014
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