Linked Life Data

2648393

Source:http://linkedlifedata.com/resource/pubmed/id/2648393

Statements in which the resource exists as a subject.
PredicateObject
rdf:type
lifeskim:mentions
pubmed:issue
7
pubmed:dateCreated
1989-5-5
pubmed:abstractText
We have developed a simple, one-step procedure for the preparation of competent Escherichia coli that uses a transformation and storage solution [TSS; 1 x TSS is LB broth containing 10% (wt/vol) polyethylene glycol, 5% (vol/vol) dimethyl sulfoxide, and 50 mM Mg2+ at pH 6.5]. Cells are mixed with an equal volume of ice-cold 2 x TSS and are immediately ready for use. Genetic transformation is equally simple: plasmid DNA is added and the cells are incubated for 5-60 min at 4 degrees C. A heat pulse is not necessary and the incubation time at 4 degrees C is not crucial, so there are no critical timing steps in the transformation procedure. Transformed bacteria are grown and selected by standard methods. Thus, this procedure eliminates the centrifugation, washing, and long-term incubation steps of current methods. Although cells taken early in the growth cycle (OD600 0.3-0.4) yield the highest transformation efficiencies (10(7)-10(8) transformants per micrograms of plasmid DNA), cells harvested at other stages in the growth cycle (including stationary phase) are capable of undergoing transformation (10(5)-10(7) transformants per micrograms of DNA). For long-term storage of competent cells, bacteria can be frozen in TSS without addition of other components. Our procedure represents a simple and convenient method for the preparation, transformation, and storage of competent bacterial cells.
pubmed:commentsCorrections
pubmed:language
eng
pubmed:journal
pubmed:citationSubset
IM
pubmed:chemical
pubmed:status
MEDLINE
pubmed:month
Apr
pubmed:issn
0027-8424
pubmed:author
pubmed:issnType
Print
pubmed:volume
86
pubmed:owner
NLM
pubmed:authorsComplete
Y
pubmed:pagination
2172-5
pubmed:dateRevised
2009-11-18
pubmed:meshHeading
pubmed:year
1989
pubmed:articleTitle
One-step preparation of competent Escherichia coli: transformation and storage of bacterial cells in the same solution.
pubmed:affiliation
Hepatitis Viruses Section, National Institute of Allergy and Infectious Diseases, Bethesda, MD 20892.
pubmed:publicationType
Journal Article, Research Support, U.S. Gov't, P.H.S., Research Support, Non-U.S. Gov't