Implantation of unmarked regulatory and metabolic modules in Gram-negative bacteria with specialised mini-transposon delivery vectors
作者： Pablo I Nikel , Víctor de Lorenzo , None
DOI: 10.1016/J.JBIOTEC.2012.05.002
关键词: Bacteria 、 Pseudomonas putida 、 Effector 、 Cell biology 、 Zymomonas mobilis 、 Genetics 、 Gram-negative bacteria 、 Transposable element 、 Escherichia coli 、 Biology 、 PEP group translocation
摘要: Engineering of robust and safe microbial cell factories requires genetic tools somewhat different from those traditionally used for laboratory-adapted microorganisms. We took advantage the properties broad-host-range mini-Tn5 vectors two regulated expression systems (LacIQ/Ptrc XylS/Pm), together with FRT-flanked, excisable antibiotic resistance determinants, to generate a set delivery gene(s) into chromosome Gram-negative bacteria. This arrangement modular elements allows cloning subsequent markerless insertion cargoes leaves behind an antibiotic-sensitive host upon action yeast Flp recombinase. engineered Pseudomonas putida KT2440 Pm::gfp strain that displayed strong fluorescence exposure 3-methylbenzoate, XylS effector, allowed us examine performance Pm promoter at single level. also reconstructed device sugar transport phosphorylation in Escherichia coli independent native phosphoenolpyruvate-dependent phosphotransferase system by stable implantation genes derived obligate anaerobe Zymomonas mobilis. In both cases, information carried implanted was stably inherited absence any selective pressure. Deliverable such as described here will enhance applicability various bacteria biocatalysis environmental bioremediation.
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