【摘 要】
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Halomonas strain TD01, a newly identified halophilic bacterium with its whole genome sequenced, proved to be a promising low-cost host for the production of
【机 构】
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CenterforQuantitativeBiologyandPeking-TsinghuaJointCenterforLifeSciences,PekingUniversity,Beijing,Ch
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Halomonas strain TD01, a newly identified halophilic bacterium with its whole genome sequenced, proved to be a promising low-cost host for the production of chemicals.However, genetic manipulation in Halomonas spp.is still difficult due to the lack of well-characterized and tunable expression systems.A method was developed enabling the efficient construction of both a constitutive promoter library and inducible promoters, porin, a highly expressed protein in Halomonas TD01, was identified using SDS-PAGE and Mass Spectrometry from its proteome.Systematic study on the upstream intergenic region of porin led to the identification of a core promoter region including-10 and-35 elements.When randomizing the sequences between-35 and-10 elements, a constitutive promoter library was constructed with 310-fold variations in the transcriptional activity;by integrating the operator of LacI into the core promoter region, an inducible promoter with a >200-fold induction enhancement was obtained.Furthermore, the constitutive and inducible promoters as two complementary expression systems were successfully employed to regulate the expression of genes for synthesizing poly-3-hydroxybutyrate (PHB) in Halomonas TD01.The method exploring the transcriptional elements shoud be able to generalize to other less-characterized bacterial strains.
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