Expression of glutathione-S-transferase fusion protein and human CCL3L1 protein / 中国医学科学院学报

ABSTRACT

<p><b>OBJECTIVE</b>To clone human CCL3L1 cDNA and to express and purify the glutathione-S-transferase (GST) fusion protein and human CCL3L1 protein.</p><p><b>METHODS</b>Total RNA was isolated from breast cancer cell line MCF7. CCL3L1 cDNA including open reading frame was obtained by RT-PCR. PCR product was digested with EcoR I and cloned into the pGEX-4T-1 vector. The plasmids from positive clone was prepared and sequenced to confirm the CCL3L1 in correct fusion form. pGEX-4T-CCL3L1 was transfected to BL21 E. coli via isopropyl-beta-D-thiogalactoside (IPTG) induction to produce GST-CCL3L1 fusion protein, which was further detected by SDS-PAGE and Western blotting.</p><p><b>RESULTS</b>As shown and confirmed by restriction endonuclease digestion analysis, CCL3L1 was correctly inserted into pGEX-4T-1 vector. The expressed fusion protein had a relative molecular weight of approximately 34 kD.</p><p><b>CONCLUSION</b>GST-CCL3L1 fusion protein can be successfully expressed using appropriate vector.</p>