摘要
目的建立Real-time PCR方法,用于定量检测核酸疫苗中宿主基因组的残留DNA。方法以Lightcycler平台为基础,选择大肠杆菌23S核糖体RNA基因为靶标基因设计扩增引物,建立基于SYBR GreenⅠ荧光染料的Real-time PCR检测方法,并用于核酸疫苗纯化过程中间产物的检测。结果整个检测过程可在30min内完成,特异性强,检测灵敏度可达10fg/μl,其标准曲线的相关系数为-0.99。结论该方法可用于核酸疫苗中宿主基因组残留DNA的检测。
Objective To develop a real-time PCR for determination of residual host genomic DNA in DNA vaccine. Methods Amplify residual host genomic DNA from DNA vaccine by PCR using designed primers target to E. coli 23S ribosome RNA gene and determine the real-time fluorescence intensity of amplified product. Determine the intermediate product during purification of DNA vaccine by the developed real-time PCR. Results The whole determination was completed within 30 min. The developed real-time PCR showed good specificity and a sensitivity of 10 fg/μl. The correlation coefficient of standard curve of the method was - 0.99. Conclusion The developed real-time PCR may be used for the determination of residual host genomic DNA in DNA vaccine.
出处
《中国生物制品学杂志》
CAS
CSCD
2007年第6期439-441,446,共4页
Chinese Journal of Biologicals
