1887

Abstract

The mevinolin-resistance determinant, , encodes the enzyme 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase and is a commonly used selectable marker in halobacterial genetics. Plasmids bearing this marker suffer from instability in because the resistance gene was derived from the genome of this species and is almost identical in sequence to the chromosomal copy. In order to reduce the level of homologous recombination between introduced plasmid vectors and the chromosome of , a homologue of the determinant was obtained from the distantly related organism, . The nucleotide sequences of the wild-type genes () of these two species are only 78% identical, and the predicted protein sequences show 71% identity. In comparison to the wild-type gene, the resistance gene from a mutant resistant to simvastatin (an analogue of mevinolin) showed a single base substitution in the putative promoter. Plasmids constructed using the new resistance determinant were stably maintained under selection in and possessed very low recombination rates with the chromosome of this species. In addition, an improved strain of was developed to overcome the plasmid instability and growth reduction observed in the commonly used WFD11 strain.

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2001-04-01
2019-11-16
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