Peer-Reviewed Journal Details
Mandatory Fields
Motherway, MO,O'Driscoll, J,Fitzgerald, GF,van Sinderen, D
2009
May
Microbial Biotechnology
Overcoming the restriction barrier to plasmid transformation and targeted mutagenesis in Bifidobacterium breve UCC2003
Validated
Optional Fields
SITE-SPECIFIC ENDONUCLEASE SEQUENCE-ANALYSIS DNA METHYLTRANSFERASES MODIFICATION SYSTEM LACTOCOCCUS-LACTIS LONGUM CONSTRUCTION STRAINS CLONING GENES
2
321
332
In silico analysis of the Bifidobacterium breve UCC2003 genome predicted two distinct loci, which encode three different restriction/modification systems, each comprising a modification methylase and a restriction endonuclease. Based on sequence homology and observed protection against restriction we conclude that the first restriction endonuclease, designated BbrI, is an isoschizomer of BbeI, the second, BbrII, is a neoschizomer of SalI, while the third, BbrIII, is an isoschizomer of PstI. Expression of each of the B. breve UCC2003 methylase-encoding genes in B. breve JCM 7017 established that BbrII and BbrIII are active and restrict incoming DNA. By exploiting knowledge on restriction/modification in B. breve UCC2003 we successfully increased the transformation efficiency to a level that allows the reliable generation of mutants by homologous recombination using a non-replicative plasmid.
10.1111/j.1751-7915.2008.00071.x
Grant Details