Liu Ming-Xue, Yin Chang-Cheng, Chun Lei, Huang Hua-Liang
Life Science and Engineering College, Southwest University of Science and Technology, Mianyang 621000, China.
Sheng Wu Gong Cheng Xue Bao. 2007 Mar;23(2):352-7.
Secondary lymphoid-tissue chemokine (SLC) is a type of CC chemokine identified by searching the Expressed Sequence Tag (EST) database. The full-length SLC gene was synthesized based on human SLC sequence using SOE-PCR. The sequenced SLC gene was cloned into expression vector pTMF and pALM, which used to transform Escherichia coli. Then the E. coli was cultured and induced according to protocol. The expressed target protein was identified by Western blotting. The target protein was expressed as soluble protein as well as inclusion bodies, the ratio of these two forms target protein varied with the difference conditions of culture and induction. The target protein was purified with the methods of nickel-nitrilotriacetic acid (Ni-NTA) metal-affinity chromatography. The results of electrophoresis of the purified target protein showed that the molecular weight was larger than the predicted molecular weight.
二级淋巴组织趋化因子（SLC）是通过搜索表达序列标签（EST）数据库鉴定出的一种CC趋化因子。基于人SLC序列，采用重叠延伸PCR（SOE-PCR）合成了全长SLC基因。将测序后的SLC基因克隆到表达载体pTMF和pALM中，用于转化大肠杆菌。然后按照方案培养和诱导大肠杆菌。通过蛋白质免疫印迹法鉴定表达的目标蛋白。目标蛋白以可溶性蛋白和包涵体的形式表达，这两种形式的目标蛋白比例随培养和诱导条件的不同而变化。采用镍-亚氨基三乙酸（Ni-NTA）金属亲和层析法纯化目标蛋白。纯化后的目标蛋白电泳结果显示，其分子量大于预测分子量。
