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目的 :研究受体与配体作用机制以及粒细胞集落刺激因子受体 (G CSFR)结合肽类似物的筛选 ,表达rG CSFR胞外CRH区。方法 :利用逆转录 聚合酶链反应 (RT PCR)技术从小鼠白血病细胞 (NFS 60 )扩增出rG CSFR胞外CRH区的cDNA片段 ,通过基因重组将该片段克隆于谷胱甘肽转移酶 (GST)融合表达载体pGEX 1λT中。结果和结论 :克隆片段与文献报道的rG CSFR序列完全一致 ,连接产物转化大肠杆菌BL 2 1,经IPTG诱导可获得大量稳定表达的CRH GST融合蛋白。
OBJECTIVE: To study the mechanism of receptor-ligand interaction and the screening of granulocyte-colony stimulating factor receptor (G CSFR) binding peptide analogs to express extracellular CRH region of rG CSFR. Methods: cDNA fragment of extracellular CRH region of rG CSFR was amplified from mouse leukemia cells (NFS 60) by reverse transcription polymerase chain reaction (RT PCR). The fragment was cloned into glutathione transferase GST) fusion expression vector pGEX 1λT. RESULTS AND CONCLUSION: The cloned fragment was identical to the rG CSFR sequence reported in the literature. The ligation product was transformed into E. coli BL21, and a large number of stably expressed CRH GST fusion proteins were obtained after IPTG induction.