Development of pyrF-based gene knockout systems for genome-wide manipulation of the archaea Haloferax mediterranei and Haloarcula hispanica

The methylaspartate cycle in haloarchaea and its possible role in carbon metabolism

“…H. hispanica strain ATCC 33960 was grown under aerobic conditions at 37°C in a 10-l flask using an air pump or in shaken 1-l flasks on a chemically defined medium (Kauri et al, 1990) with acetate (0.2%, w/v), pyruvate (0.2%) or acetate (0.2%) plus pyruvate (0.2%) as carbon sources. For the cultivation of the uracil auxotrophic (pyrF-deleted) strain DF60 (Liu et al, 2011a) and its derivatives, uracil (stock solution 50 mg ml − 1 , dissolved in DMSO) was added at a concentration of 50 mg l − 1 .…”

“…Methylaspartate cycle in haloarchaea F Borjian et al described previously (Liu et al, 2011a). The gene deletion mutants were selected by PCR verification and confirmed by sequence analysis.…”

“…Furthermore, the studies of the methylaspartate cycle in H. marismortui were hampered by the lack of a genetic system for this archaeon. Therefore, we decided to switch to H. hispanica, for which a gene deletion system is available (Liu et al, 2011a) to address these important open questions.…”

Complete Genome Sequence of the Metabolically Versatile Halophilic Archaeon Haloferax mediterranei, a Poly(3-Hydroxybutyrate- co -3-Hydroxyvalerate) Producer

“…Two key enzymes (PhaEC and PhaB) and a structure protein (PhaP) involved in PHBV biosynthesis have been identified in H. mediterranei (2,6,9,17). A highly efficient genetic manipulation system (14) and the availability of its genome sequence would expand our understanding of the relationship between PHBV biosynthesis and central metabolism in H. mediterranei. Thus, we announce the complete genome sequence of H. mediterranei CGMCC 1.2087 (ϭATCC 33500), which was obtained from the China General Microbiological Culture Collection Center (CGMCC).…”

Complete Genome Sequence of Haloarcula hispanica, a Model Haloarchaeon for Studying Genetics, Metabolism, and Virus-Host Interaction

“…It is exceptional among members of the Halobacteriaceae family in that it exhibits unusually low restriction activity and is therefore significant as one of the most tractable haloarchaea for archaeal studies (2), as well being useful as a vital host for investigations of viruses in hypersaline ecosystems (13). A transformation method for H. hispanica was established 20 years ago (2, 3), whereas the high-efficient knockout system for its genome manipulation was developed recently (11). In particular, H. hispanica has shown the ability to accumulate polyhydroxyalkanoate (PHA) when carbon sources are in excess.…”