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Query: EC:3.2.1.31 (beta-glucuronidase)
 7,680 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

A plant expression cassette was constructed using the cauliflower mosaic virus 35S 5' regulatory region with the enhancer duplicated and the 35S polyadenylation signal. Insertion of a polylinker between the transcription initiation and polyadenylation sites allows for easy cloning of genes. To test the usefulness of the cassette chimeric bacterial genes were prepared. The constructs were introduced into Nicotiana tabacum suspension culture cells by the particle bombardment process. Expression of the beta-glucuronidase reporter gene was verified by histochemical staining. Stable kanamycin and hygromycin resistant transgenic lines were obtained after introduction of chimeric genes encoding the enzymes neomycin phosphotransferase and hygromycin B phosphotransferase, respectively. The number of stable transformants was approximately 2% of the cells that transiently expressed the beta-glucuronidase reporter gene.
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PMID:The pFF plasmids: cassettes utilising CaMV sequences for expression of foreign genes in plants. 136 89

The plant-pathogenic fungus Pseudocercosporella herpotrichoides has been successfully transformed by using two different positive selection systems in combination with the Escherichia coli gusA gene. The selectable markers used in this study were the hygromycin B phosphotransferase gene (hph) from E. coli and the gene (bml) for beta-tubulin from a benomyl-resistant mutant of Neurospora crassa. A lower transformation rate was obtained with the bml system than with the hph system. Conversely, cotransformation frequencies, as determined with medium plates containing the chromogenic substrate 5-bromo-4-chloro-3-indolyl-beta-D-glucuronic acid, were higher with bml than with hph as the selectable marker. The hygromycin-resistant transformants were mitotically stable, and both the selectable gene and gusA were maintained through conidiation. The vector DNA was integrated into the genome, and the number and sites of insertion varied among transformants. Enzyme assays of mycelial extracts showed that beta-glucuronidase activity was highest in transformants with a high gusA copy number. Expression of gusA during growth of the fungus on plants was easily detectable and did not affect pathogenicity. These results form the basis for construction of a versatile and sensitive reporter gene system for P. herpotrichoides.
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PMID:Expression of the Escherichia coli beta-glucuronidase gene in Pseudocercosporella herpotrichoides. 174 51

A rapid and efficient electroporation procedure has been developed for transformation of germinating conidia of filamentous fungi. Pretreatment of conidial preparations with a cell wall weakening agent, such as beta-glucuronidase, was found to be essential for successful transformation. Using the qa-2+ gene of Neurospora crassa, encoding the catabolic dehydroquinase, as a selectable marker with a double-mutant host strain, auxotrophic for aromatic amino acids, integration of the plasmid was observed to be predominantly at ectopic chromosomal sites. Cotransformation with the qa-2+ gene and a plasmid containing a heat shock gene sequence (hsp70 of N. crassa) suggested integration site preference. High efficiencies of transformation to hygromycin resistance were achieved employing the bacterial hygromycin B phosphotransferase gene with N. crassa, the patulin-producer Penicillium urticae, and the causal agent of blackleg disease of crucifers, Leptosphaeria maculans. The economically important species Aspergillus oryzae was similarly transformed to benomyl resistance with the benomyl-resistant beta-tubulin gene of N. crassa as a dominant selectable marker.
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PMID:An electroporation-based system for high-efficiency transformation of germinated conidia of filamentous fungi. 183 30

GPD regulatory sequences were used to express a phleomycin resistance gene (Sh ble) in Schizophyllum commune, resulting in high numbers of phleomycin-resistant transformants. Attempts to express heterologous genes coding for hygromycin B phosphotransferase (hph), aminoglycoside phosphotransferase (apt), beta-glucuronidase (uidA) and beta-galactosidase (lacZ) using the same regulatory sequences were not successful and no mRNA could be detected. Cloning the hph and uidA genes in an internally deleted GPD gene resulted in truncated transcripts which ended within the 5'-parts of the heterologous genes. Cloning of the same genes as transcriptional fusions downstream from the Sh ble gene also resulted in truncated transcripts ending in the 5'-parts of these heterologous genes. It is suggested that AT-rich sequences in heterologous genes might be involved in generating these truncated transcripts, thereby preventing expression in S. commune.
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PMID:Expression of heterologous genes in Schizophyllum commune is often hampered by the formation of truncated transcripts. 950 4

The biotrophic rust fungus Puccinia graminis f. sp. tritici (Pgt) was transformed by particle bombardment. The promoter from the Pgt translation elongation factor 1alpha (EF-1alpha) gene was fused to the bacterial marker genes hygromycin B phosphotransferase (hpt) and beta-glucuronidase (GUS). Transformation constructs were introduced into uredospores of Pgt, an obligate pathogen of wheat, by biolistic bombardment. Uredospores transformed with the construct containing the hpt gene germinated and initiated branching on selective medium, indicating that they had acquired resistance to hygromycin B. However, transformants stopped growing 5 days after bombardment. GUS activity in uredospores and germlings was histochemically detected 4-16 h after bombardment. GUS expression was also obtained using the INF24 promoter from the bean rust fungus Uromyces appendiculatus, demonstrating that heterologous genes can be expressed in P. graminis under the control of regulatory sequences from closely related organisms.
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PMID:Transient transformation of the rust fungus Puccinia graminis f. sp. tritici. 1066 51

In an effort to delineate the precise mechanisms underlying the organ-specific expression of photosynthesis genes, Arabidopsis lines homozygous for each transgene construct made with the gene for hygromycin B phosphotransferase or beta-glucuronidase (GUS) placed under control of the promoter of the nuclear gene for the small subunit of ribulose-1,5-bisphosphate carboxylase/oxygenase (RBCS-3B) were constructed. Furthermore, activation tagging with T-DNA possessing quadruply repeated enhancers derived from the cauliflower mosaic virus 35S promoter was applied to a transgenic line of Arabidopsis. Mutants resistant to hygromycin B during the growth of calli generated from non-green roots on callus-inducing medium resulted from the expression of hygromycin B phosphotransferase driven by the RBCS-3B promoter. Three mutant lines, ces101 to ces103 (callus expression of RBCS), were obtained from approximately 4,000 calli resistant to a selectable marker for transformation. The active transcription driven by the RBCS-3B promoter in all the calli of ces mutants was confirmed by expression of both the GUS reporter gene and endogenous RBCS-3B. Chlorophyll and carotenoids, as well as light-dependent O(2) evolution, have been detected in the calli of all ces mutants. The loci where T-DNA was integrated in the ces101 line were determined by thermal asymmetric interlaced (TAIL)-PCR. The introduction of a DNA fragment harboring the gene for receptor-like kinase placed under the influence of enhancers into the parental line reproduced the phenotype of ces mutants. We have thus concluded that CES101 is a receptor-like kinase. The strategy presented in this investigation may promise to select a greater number of ces mutants.
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PMID:Arabidopsis mutants by activation tagging in which photosynthesis genes are expressed in dedifferentiated calli. 1659 26