The technology of large-scale pharmaceutical plasmid purification by cetyltrimethylammonium bromide and Tritonx-114

文献类型： 外文期刊

第一作者： Hou, S. H.

作者： Hou, S. H.;Zhu, H. F.;Zhang, Q.;Yuan, W. F.;Sun, H. C.

作者机构：

关键词： Plasmid;CTAB;TritonX-114;genomic DNA;RNA;endotoxin;protein;transfection

期刊名称：AFRICAN JOURNAL OF BIOTECHNOLOGY （ 影响因子：0.573； 五年影响因子：0.794 ）

ISSN： 1684-5315

年卷期： 2010 年 9 卷 1 期

页码：

收录情况： SCI

摘要： Many methods for plasmid purification have been developed, and the whole process must be designed to remove the host RNA, protein, genomic DNA and endotoxin. Currently, plasmid is mostly purified by time-consuming chromatographies. As an alternative, a new plasmid purification technology with cetyltrimethylammonium bromide (CTAB) is described. After lysis with alkali, the CTAB was directly titrated into the supernatant for plasmid precipitation, then the coprecipitated pellets were dealt with 3 M KAc and TritonX-114. Quality detection showed that the purified plasmids were free from the contamination of host RNA. In 1 mg purified plasmid, the bacterial genomic DNA, host endotoxin and protein were less than 10 mu g/mg, 50 EU/mg and 10 mu g/mg, respectively. The ratio of OD(260)/OD(280) was between 1.75 - 1.85, more than 90% of the prepared plasmid presented in the supercoiled form. Further test demonstrated that the pcDNAlacZ purified with CTAB and authoritative endotoxin-free plasmid Kit had the similar transfection efficiency in vivo and in vitro. CTAB can be used for plasmid purification; the main advantages of the DNAs purified with CTAB include the avoidance of animal-derived enzymes, toxic substance like chloroform and phenol. More attractive is that the whole process has the predominance of low cost.

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