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首页> 外文期刊>Biotechnology Progress >High-Level scu-PA Production by Butyrate-Treated Serum-Free Culture of Recombinant CHO Cell Line
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High-Level scu-PA Production by Butyrate-Treated Serum-Free Culture of Recombinant CHO Cell Line

机译：重组CHO细胞无丁酸酯处理的高浓度scu-PA生产

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摘要

The MGpUK-5 cell line,transformed with a single-chain urokinase-type plasminogen activator(scu-PA)minigene,generated mRNA transcripts and scu-PA titers corresponding to 65% or 86% of the amount generated before serum-free adaptation,despite significant loss of scu-PA gene copies during adaptation to serum-free culture.To further augment scu-PA production,a culture strategy employing sodium butyrate was explored.In 60-mL spinner flask cultures,sodium butyrate in the concentration range 1-10 mM allowed scu-PA production 2- to 3-fold higher than that in the negative control culture.Its productivity-enhancing activity was dependent on cell density in a range of 1-5X10~6 cells/mL,generating 72,200(+-)8,100 lU/mL(480(+-)50 mg/L)in 60-mL spinner flask cultures.To confirm this result,cells were grown to 4.4 x 106 cells/mL and treated with 5 mM sodium butyrate in a 2.5-L perfusion culture.The scu-PA titer increased more than 2-fold,and specific production rate of scu-PA increased 3-fold by this treatment.Overall,this perfusion culture gave rise to 1.7X10~8 IU scu-PA(1.1 g),comparable to total scu-PA production in a batch butyrate-treated culture performed at a 25-L bioreactor scale(1.3-3.5 g).Our results suggest that sodium butyrate treatment on high-density culture enables scu-PA production in gram quantities.
机译：MGpUK-5细胞系经单链尿激酶型纤溶酶原激活物（scu-PA）微型基因转化后，产生了mRNA转录本和scu-PA滴度，相当于无血清适应前产生量的65％或86％，尽管在适应无血清培养过程中scu-PA基因拷贝明显丢失。为进一步提高scu-PA的产量，探索了一种使用丁酸钠的培养策略。在60 mL旋转瓶培养物中，丁酸钠的浓度范围为1- 10 mM使scu-PA的产量比阴性对照培养物中的scu-PA高2至3倍。其生产力增强活性取决于1-5X10〜6细胞/ mL范围内的细胞密度，产生72,200（+- ）在60 mL旋转瓶培养中的8,100 lU / mL（480（+-）50 mg / L）。为证实此结果，将细胞生长至4.4 x 106细胞/ mL，并在2.5-mL溶液中用5 mM丁酸钠处理L灌注培养。据此，scu-PA滴度增加了2倍以上，而scu-PA的比生产率提高了3倍总的来说，这种灌注培养产生了1.7X10〜8 IU scu-PA（1.1 g），相当于在25 L生物反应器规模（1.3-3.5 g）下进行的丁酸分批处理培养中的总scu-PA产量。 ）。我们的结果表明，在高密度培养物中使用丁酸钠进行处理可以使scu-PA的产量达到克。

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