Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:3.2.1.23 (beta-galactosidase)
 14,648 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

A homologous transformation system for Aspergillus oryzae is described. The system is based on an A. oryzae strain deficient in orotidine-5'-phosphate decarboxylase (pyrG) and the vector pAO4-2, which contains a functional A. oryzae pyrG gene as selection marker. Transformation of the A. oryzae pyrG mutant with circular pAO4-2 resulted in the appearance of Pyr+ transformants at a frequency of up to 20 per micrograms of DNA, whereas with linear pAO4-2 up to 200 transformants per micrograms DNA were obtained. In 75% of the Pyr+ transformants recombination events had occurred at the pyrG locus, most of which (90%) resulted in insertion of one or two copies of the vector and the others (10%) in a replacement of the mutant allele by the wild-type allele. Vector pAO4-2 is also capable of transforming a corresponding mutant of Aspergillus niger. This transformation system was used to introduce into A. oryzae the heterologous and non-selectable bacterial genes lacZ, encoding beta-galactosidase, and uidA, encoding beta-glucuronidase. Using the Aspergillus nidulans gpdA promoter to drive bacterial gene expression in A. oryzae, relatively high levels of activity, as well as protein per se, as judged by western blot analyses, were obtained.
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PMID:A gene transfer system based on the homologous pyrG gene and efficient expression of bacterial genes in Aspergillus oryzae. 268 30

Expression of the URA3 gene of Saccharomyces cerevisiae was studied by analysis of URA3-lacZ gene fusions constructed in vitro. Synthesis of hybrid beta-galactosidase by fusions in frame with the coding sequence for orotidine-5'-phosphate decarboxylase (OMPdecarboxylase) was found to be normally regulated even when only 11 nucleotides of URA3 coding sequence remained, indicating that all transcription initiation and regulatory sites are present at the beginning of the URA3 gene. An upstream initiator codon that begins a short overlapping coding sequence in another reading frame was also found to be active in producing hybrid beta-galactosidase. However this beta-galactosidase synthesis showed little or no regulation. Nuclease protection experiments revealed numerous species of URA3 mRNA. The regulation of these is consistent with the idea that the URA3 protein and the overlapping peptide are translated from differentially regulated mRNAs of different lengths.
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PMID:Structure and function of the yeast URA3 gene. Differentially regulated expression of hybrid beta-galactosidase from overlapping coding sequences in yeast. 631 53

Sulfolobus solfataricus has developed into an important model organism for molecular and biochemical studies of hyperthermophilic archaea. Although a number of in vitro systems have been established for the organism, efficient tools for genetic manipulations have not yet been available for any hyperthermophile. In this work, we have developed a stable and selectable shuttle vector based on the virus SSV1 of Sulfolobus shibatae. We have introduced pUC18 for propagation in Escherichia coli and the genes pyrEF coding for orotidine-5'-monophosphate pyrophosphorylase and orotidine-5'-monophosphate decarboxylase of Sulfolobus solfataricus as selectable marker to complement pyrimidine auxotrophic mutants. Furthermore, the beta-galactosidase gene (lacS) was introduced into this vector as a reporter under the control of the strong and heat-inducible promoter of the Sulfolobus chaperonin (thermosome). After transformation of a S. solfataricus pyrEF/lacS double mutant, the vector was found to reside as a single-copy vector, stably integrated into the host chromosome via the site-specific recombination system of SSV1. Specific beta-galactosidase activities in transformants were found to be fourfold higher than in wild-type S. solfataricus cells, and increased to more than 10-fold after heat shock. Greatly increased levels of lacS mRNA were detected in Northern analyses, demonstrating that this reporter gene system is suitable for the study of regulated promoters in Sulfolobus and that the vector can also be used for the high-level expression of genes from hyperthermophilic archaea.
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PMID:A reporter gene system for the hyperthermophilic archaeon Sulfolobus solfataricus based on a selectable and integrative shuttle vector. 1278 52