摘要
通过RTPCR技术直接从激活的健康人外周血单个核细胞(HPBMC)中扩增出约03kb的hIL13基因片段,经DNA重组技术,插入到融合蛋白表达载体pGEX4T2中,转化入感受态的大肠杆菌TG1中,成功地构建了hIL13的基因工程表达菌株hIL13pGEX4T2/TG1。经异丙基硫代半乳糖苷(IPTG)诱导后表达的GSTIL13融合蛋白经SDSPAGE证明分子量约36kD,表达量约占菌体蛋白总量的10%~30%。复性后IL13融合蛋白的生物学活性可达80~120U/ml。复性后的融合蛋白经凝血酶作用后,可被切割成26kD的GST与10kD的hIL13。
The genetic engineering expression bacterial hIL13pGEX4T2/TG1 was successfully constructed through the insertion of hIL13 fragment(0.3 kb) that was directly amplified from activated healthy human peripheral blood mononuclear cells(HPBMC) by RTPCR method,into the GST fusion expression vector pGEX4T2,and the transformation into competent E.coli TG1.The molecular weight of GSTIL13 induced by IPTG is about 36 kD identified by SDSPAGE,and its expression level accounted 10%~30% of total bacterial proteins.The biological activities of renatured IL13 fusion protein could reach 80~120 U/ml and the fusion proteins could be cut into 26 kD GST and 10 kD IL13 with the treatment of thrombin for several hours.
出处
《中国免疫学杂志》
CAS
CSCD
北大核心
1998年第5期322-325,共4页
Chinese Journal of Immunology
基金
国家自然科学基金