生物技术通报 ›› 2022, Vol. 38 ›› Issue (7): 278-286.doi: 10.13560/j.cnki.biotech.bull.1985.2021-1238

• 研究报告 • 上一篇    下一篇

葡萄糖和丁酸钠对CHO-rHSA工程细胞株中rHSA产量的影响

牛宇辉1,2(), 李向茸1, 吴贝1,2, 李洪珊1,2, 李殿玉1,2, 陈磊1,2, 魏锁成2(), 冯若飞1()   

  1. 1.西北民族大学生物医学研究中心 生物工程与技术国家民委重点实验室,兰州 730030
    2.西北民族大学生命科学与工程学院,兰州 730030
  • 收稿日期:2021-09-27 出版日期:2022-07-26 发布日期:2022-08-09
  • 作者简介:牛宇辉,男,硕士研究生,研究方向:病原生物学与基因工程;E-mail: 747449152@qq.com
  • 基金资助:
    兰州市人才创新创业项目(2019-RC-20);国家民委中青年英才计划[批准号(2018)

Effects of Glucose and Sodium Butyrate on the rHSA Yield in CHO-rHSA Engineering Cell Line

NIU Yu-hui1,2(), LI Xiang-rong1, WU Bei1,2, LI Hong-shan1,2, LI Dian-yu1,2, CHEN Lei1,2, WEI Suo-cheng2(), FENG Ruo-fei1()   

  1. 1. Key Laboratory of Biotechnology & Bioengineering of State Ethnic Affairs Commission,Biomedical Research Center,Northwest Minzu University,Lanzhou 730030
    2. College of Life Science and Engineering,Northwest Minzu University,Lanzhou 730030
  • Received:2021-09-27 Published:2022-07-26 Online:2022-08-09

摘要:

目前,悬浮CHO细胞作为外源蛋白的最有潜力的工程细胞株被广泛应用,探索其高密度生长和高表达外源蛋白的培养条件也成为研究的热点。葡萄糖和丁酸钠在促进CHO细胞生长和外源蛋白表达方面具有重要作用,所以研究葡萄糖和丁酸钠对CHO-rHSA工程细胞表达重组人血清白蛋白的影响有重要的应用价值。首先利用单因素试验确定了葡萄糖和丁酸钠对CHO-rHSA工程细胞表达重组人血清白蛋白的作用,并筛选出最优添加条件;然后利用双因素组合作用试验优选出可有效提高rHSA表达量的葡萄糖及丁酸钠的组合培养条件。细胞培养第3天开始,每48 h添加终浓度为7 g/L的葡萄糖,可使rHSA产量平均提高121.93%;细胞培养第2天添加终浓度为1.0 mmoL/L的丁酸钠,可使rHSA产量平均提高110.01%;丁酸钠和葡萄糖组合使用可使CHO-rHSA工程细胞株培养时间从7 d左右延长到14 d左右,使rHSA表达量达平均达到了85.642 mg/L,rHSA表达量与未处理组相比平均提高了212.49%,比单独使用葡萄糖平均提高了134.85%,比单独使用丁酸钠平均提高了88.35%。确定待细胞培养48 h 时添加1.0 mmol/L 丁酸钠和3.0→7.0 g/L葡萄糖为CHO工程细胞株高效表达的培养条件。丁酸钠和葡萄糖组合使用可更高效的表达外源蛋白,葡萄糖可以减弱丁酸钠对CHO细胞生长的抑制作用,为提高CHO工程细胞株外源蛋白表达提供新的理论依据。

关键词: 重组人血清白蛋白, 丁酸钠, 葡萄糖, CHO-rHSA工程细胞株

Abstract:

At present,suspension CHO cells are widely used as the most potential engineered cell lines for exogenous proteins. The culture conditions of promoting its high density growth and high expression of exogenous protein have also become the research hotspot. Glucose and sodium butyrate play an important role in promoting the growth of CHO cells and the expression of exogenous proteins. Therefore,study the effect of glucose and sodium butyrate on the expression of recombinant serum albumin in CHO-rHSA engineered cells is of great value. Firstly,the effect of glucose and sodium butyrate on the expression of recombinant serum albumin in CHO-rHSA engineering cells was determined by single factor test,and the optimal addition conditions were selected. Then the combination culture conditions of glucose and sodium butyrate which effectively increased the expression of rHSA were optimized by the two-factor combination experiment. The yield of rHSA increased 121.93% by adding 7 g/L glucose every 48 h from the 3rd day of cell culture,and 110.01% by adding 1.0 mmoL/L sodium butyrate on the 2nd day of cell culture. The combination of sodium butyrate and glucose prolonged the culture time from 7 d to 14 d and increased the expression of rHSA to 85.642 mg/L,which was 212.49% higher than that of untreated cells,134.85% higher than glucose alone,and 88.35% higher than sodium butyrate alone. The culture condition of CHO engineering cell line was determined to add 1.0 mmol/L sodium butyrate and 3.0→7.0 g/L glucose when cells cultured for 48 h. Sodium butyrate combined with glucose can promote foreign protein expression more efficiently,and glucose can weaken the inhibition of sodium butyrate on the growth of CHO cells,which provides a new theoretical basis for enhancing the expression of foreign protein in CHO engineering cells.

Key words: recombinant human serum albumin,rHSA, NaBu, glucose, CHO-rHSA engineering cell line