摘要
研究了摇瓶培养条件对人α2 a干扰素酵母工程菌生长及表达的影响 ,结果表明 ,当基础料中腺嘌呤含量为 2 0μg/m L,在发酵进行至 1 2 h时再加入 2 0μg/m L腺嘌呤时 ,IFN-α2 a生物活性及比活分别达到了 7.3× 1 0 6IU/m L及 6 .77× 1 0 7IU/mg。发酵过程中后期补入适量葡萄糖 ,且维持低糖水平 ,有利表达。维持总碳源含量不变 ,使培养基中葡萄糖与蔗糖比例为 1∶ 0 .1 ,结果 IFN- α2 a生物活性及比活分别比对照增加了一倍以上。在发酵进行至 1 2 h时 ,加入 2 g/L谷氨酸 ,IFN-α2 a生物活性及比活分别比对照提高了 1 .76倍及 1 .94倍 ,而添加 0 .2 5～ 1 .0 g/L赖氨酸也能显著提高表达水平。培养基初始 p H对产物表达有较大影响 ,初始 p H为 6 .5时 ,对表达最有利。利用上述摇瓶培养优化条件 ,在 2 .6 L 生物反应器中进行补料分批培养 ,人 α2 a干扰素生物活性达到了1 .3× 1 0 7IU/m L,为原工艺的 3倍。
The effects of different culture conditions on the expression level of human IFNα2a by using recombinant yeast was investigated in shake flasks. The experimental results indicated that when 20μg/mL of adenine was added in basal culture medium, 20μg/mL of adenine was supplemented at 12h after inoculation, the antivirus activity and specific activity of IFNα2a were increased to 7.3×106IU/mL and 6.77×107IU/mg respectively. Profound expression of IFNα2a, was realized when suitable amount of glucose was added in the medium and low residue glucose concentration was kept during the midlate phase of cultivation. In addition, it was found that the biological activity and specific activity of IFNα2a were increased by more than 100% if the ratio of glucose and sucrose in the basal medium was 1∶0.1. Likewise, the addition of 2.0g/L glutamic acid at 12hfermentation time results in 176 and 194% increase of the biological activity and specific activity of IFNα2a respectively. While supplement of 0.25~1.0g/L lysine leads to the intensive enhancement of the expression level of IFNα2a. The initial pH of the basal medium proved to be very crucial to the expression level, pH 6.5 is most favovarble to the expression of the product. When the above optimal cultivation condition obtained in the shake flask was applied to the fedbatch cultivation using 2.6L jar fermenter, human IFNα2a biological activity reached 1.3×107IU/mL, which was 3 times as high as the control.
出处
《华东理工大学学报（自然科学版）》
EI
CAS
CSCD
北大核心
2000年第3期279-283,共5页
Journal of East China University of Science and Technology
关键词
α2a型干扰素
发酵
培养条件
酵母工程菌
Saccharomyces cerevisiae
cultivation
recombinant yeast
IFNα2a
adenine
