Rapid preparation of total nucleic acids from E. coli for multi-purpose applications.
Separate protocols are commonly used to prepare plasmid DNA, chromosomal DNA, or total RNA from E. coli cells. Various methods for the rapid preparation of plasmid DNA have been developed previously, but the preparation of the chromosomal DNA and total RNA are usually laborious. We report here a simple, fast, reliable, and cost-effective method to extract total nucleic acids from E. coli by direct lysis of the cells with phenol. Five distinct and sharp bands, which correspond to chromosomal DNA, plasmid DNA, 23S rRNA, 16S rRNA, and a mixture of small RNA, were observed when analyzing the prepared total nucleic acids on a regular 1-2% agarose gel. The simple and high-quality preparation of the total nucleic acids in a single tube allowed us to rapidly screen the recombinant plasmid, as well as to simultaneously monitor the change of the plasmid copy number and rRNA levels during the growth of E. coli in the liquid medium.
Department of Biotechnology, College of Life Sciences, Wuhan University, Wuhan, Hubei 439972, China.
Lin
L
Cheng
Tai-Yuan
TY
Li
Yi
Y
Zhang
D001681:Biological Assay / Q000191:economics / Q000379:methods*; D004926:Escherichia coli / Q000235:genetics* / Q000502:physiology; D009696:Nucleic Acids / Q000302:isolation & purification*; D019800:Phenol / Q000378:metabolism; D010957:Plasmids / Q000235:genetics / Q000378:metabolism; D013997:Time Factors
D009696:Nucleic Acids; D019800:Phenol