[Construction of Streptococcus mutans surface protein antigen eukaryotic expression vector pcDNA3-PAc. II. Testified efficiency of repeated using glass fiber chromatographic column].
OBJECTIVE
To explore one method getting highly purified plasmid DNA and costing low.


METHODS
According to principle of extraction and purification plasmid DNA of glass fiber chromatographic column, its operation sequence was improved so that disposable glass fiber chromatographic column could be used repeatedly. The concentration and purity of plasmid DNA obtained by 12 times repeated using glass fiber chromatographic column were determined by spectrophotometry. Moreover, Those plasmid DNA obtained were tested by restriction enzyme and agarose gel electrophoresis, Ligation in vitro and transformation test.


RESULTS
12 times repeated use of glass fiber chromatographic column made no significant difference to the concentration and purity of obtained plasmid DNA (P < 0.05), and plasmid DNA was completely digested by restriction enzyme. The digested fragments were well ligated together by the action of T4 DNA Ligase in ATP ligating buffer in vitro and ligated DNA was well transformed into E. coli.


CONCLUSIONS
Repeated use glass fiber chromatographic column can obtain a lot of highly purified plasmid DNA and cost is low. Digestion, ligation and transformation of those plasmid DNA are not affected. The results may be significant in molecular cloning and study of gene vaccination.
Liu
J
J
College of Stomatology, West China University of Medical Sciences.
Liu
T
T
Tan
H
H
Zhou
X
X
Huang
X
X
Guo
L
L
D000954:Antigens, Surface; D002855:Chromatography, Thin Layer / Q000379:methods*; D004269:DNA, Bacterial / Q000302:isolation & purification*; D004926:Escherichia coli / Q000235:genetics; D005057:Eukaryotic Cells; D010957:Plasmids / Q000235:genetics*; D013295:Streptococcus mutans / Q000235:genetics*
D000954:Antigens, Surface; D004269:DNA, Bacterial