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目的 建立纤溶酶原饼环区 5 (Humanplasminogenkringle 5 ,hPK 5 )蛋白重组大肠杆菌高效表达体系 ,为高密度发酵创造条件。方法 观察一级、二级种子的生长状态 ,比较表达hPK 5蛋白的 4种重组工程菌株在相同条件下的表达情况 ,选出首选发酵种子 ;对其培养时间、诱导时间、培养基种类、pH等条件进行优化 ;并用凝胶成像分析系统对SDS PAGE结果进行分析。结果 经过筛选 ,JM10 9 pBV2 2 0 hPK 5 (简称JP5 )是获取hPK 5蛋白的首选工程菌株 ,其最佳表达条件是LB培养基 (pH 7.4、溶解氧充足 )、30℃培养 3h、4 2℃诱导 6h。在此条件下 ,JP5表达目的蛋白占菌体总蛋白的 38%左右。结论 为高密度发酵获取hPK 5蛋白奠定了实验基础
OBJECTIVE To establish an efficient expression system of recombinant plasmids of humanplasminogenkringle 5 (hPK 5) protein and to create the conditions for high-density fermentation. Methods The growth status of primary and secondary seeds was observed. The expression of the four recombinant strains expressing hPK 5 protein was compared under the same conditions, and the preferred fermentation seeds were selected. The culture time, induction time, medium type, pH And other conditions to optimize; and gel imaging analysis system SDS PAGE results were analyzed. Results After screening, JM109 pBV2 20 hPK 5 (referred to as JP5) was the first choice strain for obtaining hPK 5 protein. The optimal conditions for the expression of hPK 5 protein were LB medium (pH 7.4, sufficient dissolved oxygen), 3 h incubation at 30 ℃, 42 ℃ induced 6h. Under this condition, the expressed protein of JP5 accounted for about 38% of the total bacterial proteins. Conclusion The experimental basis for obtaining hPK 5 protein by high density fermentation was established