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  • Gene cartridges  (1)
  • Key words: diel patterns, transcription, rbcL, cyanobacterium, prymnesiophyte.  (1)
  • 1
    ISSN: 1432-0983
    Keywords: Saccharomyces cerevisiae ; G418 resistance ; Gene cartridges ; Heterologous Gene expression
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary Coding sequence cartridges for aminoglycoside phosphotransferase (APT) were isolated from bacterial transposon Tn903. When incorporated into a heterologous gene construction utilising the PGK1 promoter and terminator, the heterologous APT gene provided a G418-resistance determinant that functioned efficiently as a dominant marker for yeast in both multiple- and single-copy. Transformant colonies on selective medium appeared rapidly, within 36–48 h, and growth rate of the transformed cells was normal. A simple and highly sensitive radiolabelling assay for APT enzyme activity was developed for use with crude cell protein extracts. Enzyme activity units were equated to the amount of APT protein present in the cells, and the APT protein was shown to be stable in yeast. Heterologous APT expression was 130-fold reduced compared with homologous PGK1. This resulted from an estimated two-fold decrease in mRNA level and a 65-fold decrease in translation efficiency. The latter was unaffected by AUG sequence context change, but corresponded with a high frequency of minor codons in the APT-coding sequence. APT can be used as a semi-quantitative reporter of gene expression, whose useful features are in vivo detection via the G418-resistance phenotype and powerful cell-free assay.
    Type of Medium: Electronic Resource
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  • 2
    Electronic Resource
    Electronic Resource
    Springer
    Marine biotechnology 2 (2000), S. 429-436 
    ISSN: 1436-2236
    Keywords: Key words: diel patterns, transcription, rbcL, cyanobacterium, prymnesiophyte.
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Abstract: Diel patterns of rbcL transcription, ribulose-1,5-bisphosphate carboxylase/oxygenase (RubisCO) activity, and whole cell carbon fixation were compared in the marine cyanobacterium Synechococcus PCC7002 and the chromophytic prymnesiophyte Pavlova gyrans. Both organisms were grown on a 12:12 light-dark cycle, with the light period starting at 0700. Strong diel patterns in these three aspects of carbon fixation occurred in both organisms, with maximal levels in the light period and minima in the dark. In Synechococcus, maximal rbcL transcript abundance occurred at noon and was followed by rapid disappearance. RubisCO enzyme activity and whole cell carbon fixation were elevated at 1600, and they disappeared over the next 8 hours. In contrast, in Pavlova, rbcL transcript abundance was maximal at 1600, and it was maintained at 66% of this level into the dark period (2000). Whole cell carbon fixation and RubisCO activity were elevated into the dark period (at 2000), being 77% and 81%, respectively, of the maximum. A similar diel pattern of cyanobacterial-like and chromophyte-like rbcL transcription has been observed in natural phytoplankton populations. These studies suggest that chromophytes are more adapted to take advantage of carbon fixation late in the day than cyanophytes.
    Type of Medium: Electronic Resource
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