Linked Life Data

12406220

Source:http://linkedlifedata.com/resource/pubmed/id/12406220

Statements in which the resource exists as a subject.
PredicateObject
rdf:type
lifeskim:mentions
pubmed:issue
2
pubmed:dateCreated
2002-10-30
pubmed:databankReference
pubmed:abstractText
Progress towards understanding the molecular basis of virulence in Clostridium difficile has been hindered by the lack of effective gene transfer systems. We have now, for the first time, developed procedures that may be used to introduce autonomously replicating vectors into this organism through their conjugative, oriT-based mobilization from Escherichia coli donors. Successful transfer was achieved through the use of a plasmid replicon isolated from an indigenous C. difficile plasmid, pCD6, and through the characterization and subsequent circumvention of host restriction/modification (RM) systems. The characterized replicon is the first C. difficile plasmid replicon to be sequenced and encodes a large replication protein (RepA) and a repetitive region composed of a 35 bp iteron sequence repeated seven times. Strain CD6 has two RM systems, CdiCD6I/M.CdiCD6I and CdiCD6II/M. CdiCD6II, with equivalent specificities to Sau96I/M. Sau96I (5'-GGNMCC-3') and MboI/M. MboI (5'-GMATC-3') respectively. A second strain (CD3) possesses a type IIs restriction enzyme, Cdi I, which cleaves the sequence 5'-CATCG-3' between the fourth and fifth nucleotide to give a blunt-ended fragment. This is the first time that an enzyme with this specificity has been reported. The sequential addition of this site to vectors showed that each site caused between a five- and 16-fold reduction in transfer efficiency. The transfer efficiencies achieved with both strains equated to between 1.0 x 10-6 and 5.5 x 10-5 transconjugants per donor.
pubmed:language
eng
pubmed:journal
pubmed:citationSubset
IM
pubmed:chemical
pubmed:status
MEDLINE
pubmed:month
Oct
pubmed:issn
0950-382X
pubmed:author
pubmed:issnType
Print
pubmed:volume
46
pubmed:owner
NLM
pubmed:authorsComplete
Y
pubmed:pagination
439-52
pubmed:dateRevised
2006-11-15
pubmed:meshHeading
pubmed:year
2002
pubmed:articleTitle
Conjugative transfer of clostridial shuttle vectors from Escherichia coli to Clostridium difficile through circumvention of the restriction barrier.
pubmed:affiliation
Research Division, Centre for Applied Microbiology and Research, Wiltshire, UK.
pubmed:publicationType
Journal Article, Research Support, Non-U.S. Gov't, Evaluation Studies