Open AccessScreening costramid libraries for chromosomal genes: an alternative interspecific hybridization method
Author(s) -
Stanley J.,
Dowling D.N.,
Stucker M.,
Broughton W.J.
Publication year - 1987
Publication title -
fems microbiology letters
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.899
H-Index - 151
eISSN - 1574-6968
pISSN - 0378-1097
DOI - 10.1111/j.1574-6968.1987.tb02509.x
Subject(s) - biology , cosmid , gene , genetics , suppression subtractive hybridization , homology (biology) , genomic library , escherichia coli , gene expression , peptide sequence , cdna library
Broad host‐range RK2‐based cosmid vectors (‘costramids’) are increasingly used in molecular genetic studies of Gram‐negative soil bacteria such as Rhizobium spp. we describe a simple modification of existing methods, whereby a genomic library constructed in a stringently replicated vector can be screened for genes which are undetectable by colony hybridization due to background cross‐hybridization. This method allows the use of ‘heterologous’ probes (interspecies hybridization) to isolate several presumptive genes of interest from a gene bank of Rhizobium sp. NGR234 made in the costramid pRK7813. These are a gene with homology to the citrate synthase gene ( gltA ) or Escherichia coli , the gene encoding δ‐aminol evulinic acid synthase ( hemA ), and a gene or genes regulating dicarboxylate transport.