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  • picornavirus  (2)
  • Cloning  (1)
  • Springer  (3)
  • 1
    Electronic Resource
    Electronic Resource
    Springer
    Molecular genetics and genomics 214 (1988), S. 158-161 
    ISSN: 1617-4623
    Keywords: Vitreoscilla ; Hemoglobin ; Cloning ; Nucleotide sequence
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary Vitreoscilla hemoglobin is involved in oxygen metabolism of this bacterium, possibly in an unusual role for a microbe. We have isolated the Vitreoscilla hemoglobin structural gene from a pUC19 genomic library using mixed oligodeoxy-nucleotide probes based on the reported amino acid sequence of the protein. The gene is expressed in Escherichia coli from its natural promoter as a major cellular protein. The nucleotide sequence, which is in complete agrecment with the known amino acid sequence of the protein, suggests the existence of promoter and ribosome binding sites with a high degree of homology to consensus E. coli upstream sequences. In the case of at least some amino acids, a codon usage bias can be detected which is different from the biased codon usage pattern in E. coli. The down-stream sequence exhibits homology with the 3′ end sequences of several plant leghemoglobin genes. E. coli cells expressing the gene contain greater than fivefold more heme than controls.
    Type of Medium: Electronic Resource
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  • 2
    Electronic Resource
    Electronic Resource
    Springer
    Cytotechnology 28 (1998), S. 111-126 
    ISSN: 1573-0778
    Keywords: autoregulation ; cell-cycle engineering ; eukaryotic operon ; IRES ; multigene engineering ; picornavirus ; pTRIDENT ; regulated expression
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Abstract Contemporary basic research is rapidly revealing increasingly complex molecular regulatory networks which are often interconnected via key signal integrators. These connections among regulatory and catalytic networks often frustrate bioengineers as promising metabolic engineering strategies are bypassed by compensatory metabolic responses or cause unexpected, undesired outcomes such as apoptosis, product protein degradation or inappropriate post- translational modification. Therefore, for metabolic engineering to achieve greater success in mammalian cell culture processes and to become important for future applications such as gene therapy and tissue engineering, this technology must be enhanced to allow simultaneous, in cases conditional, reshaping of metabolic pathways to access difficult-to-attain cell states. Recent advances in this new territory of multigene metabolic engineering are intimately linked to the development of multicistronic expression technology which allows the simultaneous, and in some cases, regulated expression of several genes in mammalian cells. Here we review recent achievements in multicistronic expression technology in view of multigene metabolic engineering.
    Type of Medium: Electronic Resource
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  • 3
    ISSN: 1573-0778
    Keywords: CITE ; EMCV ; GFP ; IRES ; picornavirus
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Abstract Based on internal ribosomal entry sites (IRES) of picornaviral origin we constructed a novel family of mammalian expression vectors. pQuattro vectors contain quattrocistronic artificial eukaryotic operons which link, in a single transcript, the simultaneous and coordinated as well as adjustable expression of up to three independent genes of interest to a terminal neomycin (neo) resistance marker. Due to the strict genetic linkage of the transgenes and the terminal selection marker, this genetic configuration enables, by the selection on neomycin, multigene metabolic engineering of mammalian cells in a single step (one-step metabolic engineering). Furthermore, selection on the terminal cistron of multicistronic expression units enforces cocistronic expression of all upstream encoded genes and maximises genetic integrity of the eukaryotic operon in stable mammalian cell lines, since clones harbouring damaged multicistronic expression units become neomycin-sensitive and are automatically counterselected (auto-selection). The modular set-up and the abundance of restriction sites in pQuattro vectors facilitate the movement of individual genes between multicistronic expression vectors and guarantees high compatibility with genetic elements of a wide variety of existing mammalian expression vectors.
    Type of Medium: Electronic Resource
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