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实时定量PCR法检测生物技术药物中宿主基因组DNA残留
引用本文:曹晨华,刘晓志,段月娇,刘素霞,赵伟,常亮,纪丽曼,高健. 实时定量PCR法检测生物技术药物中宿主基因组DNA残留[J]. 生物技术进展, 2014, 4(2): 142-145. DOI: 10.3969/j.issn.2095-2341.2014.02.12
作者姓名:曹晨华  刘晓志  段月娇  刘素霞  赵伟  常亮  纪丽曼  高健
作者单位: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
基金项目:国家863 计划项目(2012AA02A306);国家科技重大专项项目(2012ZX09101313)资助。
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  
收稿时间:2013-12-13

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. Detecting Residual Host Genomic DNA in Biotech Drugs by Real-Time Quantitative PCR[J]. CURRENT BIOTECHNOLOGY, 2014, 4(2): 142-145. DOI: 10.3969/j.issn.2095-2341.2014.02.12
Authors:CAO Chen-hua§  LIU Xiao-zhi§  DUAN Yue-jiao  LIU Su-xia  ZHAO Wei  CHANG Liang  JI Li-man  GAO Jian
Affiliation:State Key Laboratory of Antibody Research and Development; New Drug Research and Development Company Ltd., North China Pharmaceutical Corporation, Shijiazhuang 050015, China
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.
Keywords:real-time quantitative PCR  residual host genomic DNA  biotech drugs  quality control  
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