Large-scale purification of pharmaceutical-grade plasmid DNA using tangential flow filtration and multi-step chromatography
2013
Sun, Bo | Yu, Xianghui | Yin, Yuhe | Liu, Xintao | Wu, Yongge | Chen, Yan | Zhang, Xizhen | Jiang, Chunlai | Kong, Wei
The demand for pharmaceutical-grade plasmid DNA in vaccine applications and gene therapy has been increasing in recent years. In the present study, a process consisting of alkaline lysis, tangential flow filtration, purification by anion exchange chromatography, hydrophobic interaction chromatography and size exclusion chromatography was developed. The final product met the requirements for pharmaceutical-grade plasmid DNA. The chromosomal DNA content was <1 μg/mg plasmid DNA, and RNA was not detectable by agarose gel electrophoresis. Moreover, the protein content was <2 μg/mg plasmid DNA, and the endotoxin content was <10 EU/mg plasmid DNA. The process was scaled up to yield 800 mg of pharmaceutical-grade plasmid DNA from approximately 2 kg of bacterial cell paste. The overall yield of the final plasmid DNA reached 48%. Therefore, we have established a rapid and efficient production process for pharmaceutical-grade plasmid DNA.
اظهر المزيد [+] اقل [-]الكلمات المفتاحية الخاصة بالمكنز الزراعي (أجروفوك)
المعلومات البيبليوغرافية
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