趋化因子配体18重组成熟肽段的表达

目的构建PET SUMO-CCL18原核表达质粒，并表达、纯化获得重组趋化因子配体18成熟肽段。方法从人卵巢癌组织中克隆CCL18基因全长，继而扩增表达成熟蛋白质的核酸序列，连接人PETSUMO载体，重组阳性克隆转入感受态细胞BL21（DE3）中IPTG诱导表达，基质辅助激光解吸／电离质谱技术（MALDI-TOF）验证后以SUMO蛋白酶切除载体部分，纯化浓缩，MALDI-TOF鉴定表达结果。结果测序分析证实，克隆人PETSUMO载体的CCL18序列与Genbank中报道序列完全一致，纯化后蛋白质谱鉴定与天然CCL18成熟蛋白序列一致。结论成功构建了高表达CCL18蛋白的表达系统，得到重组CCL18成熟肽段，为进一步研究CCL18蛋白在卵巢癌中的作用提供实验基础。

Expression of Recombinant Chemokine Ligand 18 Mature Peptide

Objective To construct PET SUMO-CCL18. prokaryotic expression plasmid, express, purify and obtain recombinant CCL18 mature peptide. Methods The DNA fragment of chemokine ligand 18 （CCL18） expressing the mature protein was cloned from human ovarian cancer, and inserted into PET SUMO vector to construct the PET SUMO-CCL18 prokaryotic expression plasmid. The positive recombinant plasmid was transformed into BI21（DE3） competent cells, and then induced with IPTG. Expression of recombinant CCL18 was identified by SDS-PAGE and matrix-assisted laser desorption/ionization mass spectrometry （MALDI-TOF） The vector protein was removed using SUMO protease, and then The recombinant CCL18 mature peptide purified with Ni-chelation affinity chromatography column. was identified and analyzed by MALDI-TOF again. Results Sequence analysis confirmed that the sequence of the recombinant CCL18 inserted in to PET SUMO is completely identical to that of Genbank as reported. Also, MALDI-TOF identified the identity of the purified recombinant CCL18 mature peptide to nature CCL18 mature protein. Conclusions The constructed prokaryotic expression system of PET SUMO-CCL18 containing recombinant CCL18 mature peptide may provide an experimental tool for further study of ovarian cancer.