Linked Life Data

1660428

Source:http://linkedlifedata.com/resource/pubmed/id/1660428

Statements in which the resource exists as a subject.
PredicateObject
rdf:type
lifeskim:mentions
pubmed:issue
1
pubmed:dateCreated
1992-1-13
pubmed:abstractText
We have developed a versatile method for integration of modified genes and gene fusions into the bacteriophage lambda attachment site (attB) of the Escherichia coli chromosome. The method relies on two components: (1) a DNA integration cassette, flanked by multiple restriction enzyme sites, which contains the lambda attP site and, as a selectable marker, the Tn5 aphA gene conferring kanamycin resistance (KmR); and (2) a plasmid with the lambda int gene transcribed from the tet promoter. A fragment carrying the gene in question is ligated to the integration cassette, resulting in a circular piece of DNA unable to replicate. The ligation product is then transformed into a strain that contains the int-carrying plasmid. Selection for KmR results in colonies with the cassette integrated into the attB site of the E. coli chromosome. This method was used for integration of several lacZ and phoA promoter fusions. The integration products were analyzed by Southern hybridization. In addition, we found, fortuitously, that the ligated DNA circles could also integrate by homologous recombination, although usually at a much lower frequency than the Int-mediated integration into attB.
pubmed:language
eng
pubmed:journal
pubmed:citationSubset
IM
pubmed:chemical
pubmed:status
MEDLINE
pubmed:month
Oct
pubmed:issn
0378-1119
pubmed:author
pubmed:issnType
Print
pubmed:day
30
pubmed:volume
107
pubmed:geneSymbol
attP, dnaA, int, lacZ, phoA
pubmed:owner
NLM
pubmed:authorsComplete
Y
pubmed:pagination
11-7
pubmed:dateRevised
2008-11-21
pubmed:meshHeading
pubmed:year
1991
pubmed:articleTitle
A versatile method for integration of genes and gene fusions into the lambda attachment site of Escherichia coli.
pubmed:affiliation
Department of Microbiology, Technical University of Denmark, Lyngby.
pubmed:publicationType
Journal Article, Research Support, Non-U.S. Gov't