Abstract:
:A spontaneous rpsL mutant of Thermus thermophilus was isolated in a search for new selection markers for this organism. This new allele, named rpsL1, encodes a K47R/K57E double mutant S12 ribosomal protein that confers a streptomycin-dependent (SD) phenotype to T. thermophilus. Models built on the available three-dimensional structures of the 30S ribosomal subunit revealed that the K47R mutation directly affects the streptomycin binding site on S12, whereas the K57E does not apparently affect this binding site. Either of the two mutations conferred the SD phenotype individually. The presence of the rpsL1 allele, either as a single copy inserted into the chromosome as part of suicide plasmids or in multicopy as replicative plasmids, produced a dominant SD phenotype despite the presence of a wild-type rpsL gene in a host strain. This dominant character allowed us to use the rpsL1 allele not only for positive selection of plasmids to complement a kanamycin-resistant mutant strain, but also more specifically for the isolation of deletion mutants through a single step of negative selection on streptomycin-free growth medium.
journal_name
Appl Environ Microbioljournal_title
Applied and environmental microbiologyauthors
Blas-Galindo E,Cava F,López-Viñas E,Mendieta J,Berenguer Jdoi
10.1128/AEM.00751-07subject
Has Abstractpub_date
2007-08-01 00:00:00pages
5138-45issue
16eissn
0099-2240issn
1098-5336pii
AEM.00751-07journal_volume
73pub_type
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