Curr. Biotech. ›› 2014, Vol. 4 ›› Issue (2): 142-145.DOI: 10.3969/j.issn.2095-2341.2014.02.12

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Detecting Residual Host Genomic DNA in Biotech Drugs by Real-Time Quantitative PCR

CAO Chen-hua, LIU Xiao-zhi, DUAN Yue-jiao, LIU Su-xia, ZHAO Wei, CHANG Liang, JI Li-man, GAO Jian   

  1. State Key Laboratory of Antibody Research and Development; New Drug Research and Development Company Ltd., North China Pharmaceutical Corporation, Shijiazhuang 050015, China
  • Received:2013-12-13 Online:2014-03-25 Published:2014-03-03

实时定量PCR法检测生物技术药物中宿主基因组DNA残留

曹晨华§,刘晓志§,段月娇,刘素霞,赵伟,常亮,纪丽曼*,高健*   

  1. 华北制药集团新药研究开发有限责任公司; 抗体药物研制国家重点实验室, 石家庄 050015
  • 通讯作者: 纪丽曼，高级工程师，从事蛋白药物研发。高健，高级工程师，从事蛋白药物研发。
  • 作者简介:§曹晨华与刘晓志为本文共同第一作者。曹晨华，工程师，从事抗体类药物工艺开发研究。E-mail: chenhuacao@hotmail. com。刘晓志，高级工程师，从事蛋白药物研发。
  • 基金资助:

    国家863 计划项目(2012AA02A306)；国家科技重大专项项目(2012ZX09101313)资助。

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Abstract: We detected residual host DNA in yeast expressing biotech drugs by SYBR GreenⅠ fluorescent based real-time quantitative polymerase chain reaction. The detection sensitivity of this method was up to 1.0 fg/μL. The linear correlation coefficient of standard curve was above 0.99 when DNA concentration was bwtween 1.0 fg/μL to 1.0 ng/μL. Three batch of samples was determined, and the residual host DNA were 8.635×105 fg/μL、6.265×102 fg/μL and 1.436  fg/μL respectively. The results suggested that this easy-handled, rapid and highly sensitive method could be used for the determination of residual yeast DNA in biotech drug products.

Key words: real-time quantitative PCR, residual host genomic DNA, biotech drugs, quality control

摘要： 利用基于SYBR GreenⅠ荧光染料的实时定量PCR方法检测酵母表达生物技术药物产品中宿主DNA残留量。该方法检测灵敏度可达到1.0 fg/μL， DNA浓度在1.0 fg/μL～1.0 ng/μL范围内线性良好，其标准曲线的相关系数为099以上。应用该方法对3批不同实验样本进行测定，宿主DNA残留量分别为8.635×105 fg/μL、6.265×102 fg/μL和1436 fg/μL 。实验表明该方法操作简便、灵敏度高，可用于生物技术药物产品中酵母DNA残留的定量测定。

关键词: 实时定量PCR, 宿主基因组DNA残留, 生物技术药物, 质量控制

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