Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: 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)

This chapter summarizes four powerful assays for analyzing gene expression in cis-regulatory studies. The enzymatic assays (CAT, luciferase, lacZ) are currently limited by their application to embryo homogenates or fixed samples, but offer more robust analysis of gene activity than GFP. Assays based on CAT enzymatic activity or on CAT mRNA detection by WMISH are laborious but are well established for accurately quantifying gene expression and to determine spatial patterns at defined timepoints during development. LacZ assays are the current standard for spatially visualizing gene products in whole-mount fixed embryos. They are very sensitive but they provide limited temporal or quantitative information due to the perdurance of beta-galactosidase and the subtleties of the staining technique. Recently developed luciferase assays promise to be even more sensitive and accurate than the CAT and lacZ assays, and applicable to living cells and embryos. But, they have not yet been well established in invertebrate deuterostome research. GFP allows visualization of gene expression within living embryos. But because this is not an enzymatic assay, sensitivity can be a problem, particularly for weak promoters. Furthermore, imaging live embryos and quantifying gene expression in space and time (due to scattering of light by tissue, the perdurance of GFP, and other experimental details) is currently fraught with challenges. Ongoing improvements in imaging technology and the advent of multiple fluorescent proteins, as well as fluorescent and luminescent assays for vital imaging, will dramatically facilitate studies of gene expression in the coming decade.
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PMID:Using reporter genes to study cis-regulatory elements. 1557 24

The taxonomic position of three novel sea-water isolates was determined. The strains studied were strictly aerobic, heterotrophic, pigmented, motile by gliding, Gram-negative and oxidase-, catalase-, beta-galactosidase- and alkaline phosphatase-positive. 16S rRNA gene sequence phylogenetic analysis indicated that the strains KMM 6020T, KMM 6021 and KMM 6028 occupied a distinct lineage within the family Flavobacteriaceae. The major respiratory quinone was MK-6. The predominant fatty acids were i15 : 0, i15 : 1, i15 : 0 3-OH, i17 : 1omega9c and i17 : 0 3-OH. On the basis of phenotypic, chemotaxonomic, genotypic and phylogenetic characteristics, the novel bacteria were assigned to the genus Aquimarina gen. nov., as Aquimarina muelleri gen. nov., sp. nov. The type strain is KMM 6020T (=KCTC 12285T=LMG 22569T). From the results of the 16S rRNA gene sequence analysis and phenotypic features, the species [Cytophaga] latercula Lewin 1969 is proposed to be reclassified in the new genus Stanierella as Stanierella latercula gen. nov., comb. nov., with type strain CIP 104806T (=ATCC 23177T=NCIMB 1399T=LMG 1343T).
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PMID:Description of Aquimarina muelleri gen. nov., sp. nov., and proposal of the reclassification of [Cytophaga] latercula Lewin 1969 as Stanierella latercula gen. nov., comb. nov. 1565 78

The taxonomic position of a novel marine bacterium isolated from the green alga Ulva fenestrata collected in the Sea of Japan was established. Cells of the strain studied, designated KMM 6017T, were strictly aerobic, heterotrophic, pink-pigmented, non-motile by gliding, Gram-negative and oxidase-, catalase-, beta-galactosidase- and alkaline phosphatase-positive. 16S rRNA gene sequence analysis indicated that the strain occupied a distinct lineage within the phylum 'Bacteroidetes' and formed a cluster with [Flexibacter] tractuosus and Reichenbachia agariperforans. The G+C content of the DNA of KMM 6017T was 40.2 mol%. The major respiratory quinone was MK-7. The predominant fatty acids were i15 : 1, i15 : 0 and i17 : 0 3-OH (34.2, 24 and 7.7 %, respectively). On the basis of phenotypic, chemotaxonomic, genotypic and phylogenetic characteristics, the novel bacterium was assigned to the genus Roseivirga gen. nov., as Roseivirga ehrenbergii gen. nov., sp. nov. The type strain is KMM 6017T (=KCTC 12282T=LMG 22567T).
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PMID:Roseivirga ehrenbergii gen. nov., sp. nov., a novel marine bacterium of the phylum 'Bacteroidetes', isolated from the green alga Ulva fenestrata. 1565 79

A bacterial strain, designated KMM 6049T, was isolated from the sea urchin Strongylocentrotus intermedius inhabiting the Sea of Japan. The bacterium studied was strictly aerobic, heterotrophic, yellow-pigmented, non-motile, Gram-negative and oxidase-, catalase-, beta-galactosidase- and alkaline phosphatase-positive. 16S rRNA gene sequence analysis indicated that strain KMM 3524T was closely related to Salegentibacter holothuriorum and Salegentibacter salegens (sharing 97.7 and 98 % sequence similarity, respectively). DNA-DNA relatedness levels between strains KMM 6049T and S. holothuriorum KMM 3524T and S. salegens DSM 5424T were 24 and 45 %, respectively, indicating that KMM 6049T belongs to a novel species of the genus Salegentibacter, for which the name Salegentibacter mishustinae sp. nov. is proposed. The type strain is KMM 6049T (=KCTC 12263T=LMG 22584T=NBRC 100592T).
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PMID:Salegentibacter mishustinae sp. nov., isolated from the sea urchin Strongylocentrotus intermedius. 1565 80

A novel marine bacterium, strain KMM 6050T, was isolated from the sea urchin Strongylocentrotus intermedius, which inhabits the Sea of Japan. The strain studied was strictly aerobic, heterotrophic, yellow-orange-pigmented, motile by gliding, Gram-negative and oxidase-, catalase-, beta-galactosidase- and alkaline phosphatase-positive. The results of 16S rRNA gene sequence analysis showed that strain KMM 6050T occupies a distinct lineage within the family Flavobacteriaceae and is most closely related to the species Mesonia algae and Salegentibacter salegens (sequence similarity of 92.5-92.6 %). The DNA G+C content of KMM 6050T was 39.6 mol%. The major respiratory quinone was MK-6. The predominant fatty acids were i15 : 0, a15 : 0, 15 : 0, i16 : 1, i16 : 0, i16 : 0 3-OH and i17 : 0 3-OH. On the basis of phenotypic, chemotaxonomic, genotypic and phylogenetic characteristics, the novel bacterium has been assigned to the genus Gramella gen. nov., as Gramella echinicola sp. nov. The type strain is KMM 6050T (=KCTC 12278T=NBRC 100593T=LMG 22585T).
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PMID:Gramella echinicola gen. nov., sp. nov., a novel halophilic bacterium of the family Flavobacteriaceae isolated from the sea urchin Strongylocentrotus intermedius. 1565 6

To facilitate gene expression analysis in the human gastric pathogen Helicobacter pylori, we constructed the plasmids pHPLAC-KAN and pHPLAC-CAT containing a promoterless Escherichia coli lacZ gene located upstream from the antibiotic resistance genes aphA-3 or cat, respectively. The suitability of the plasmids for H. pylori mutagenesis and gene expression analysis was evaluated by plasmid integration into the genome of H. pylori strain 1061 by single homologous recombination, using the rpl9 gene encoding ribosomal protein L9 as target. By monitoring beta-galactosidase production from the resulting rpl9::lacZ fusion, it was demonstrated that H. pylori rpl9 displays the classical growth phase-dependent regulation of components of the protein synthesis machinery, as beta-galactosidase production dropped fivefold in the stationary growth phase. The plasmids described in this study extend our methodological repertoire for genetic modification and molecular analysis of H. pylori, and may also be of use for other bacteria, as the resistance cassettes and the lacZ gene are active in the related Campylobacter species.
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PMID:Novel plasmids for gene expression analysis and for genetic manipulation in the gastric pathogen Helicobacter pylori. 1586 10

Cell-free protein synthesis reactions have not been seriously considered as a viable method for commercial protein production mainly because of high reagent costs and a lack of scalable technologies. Here we address the first issue by presenting a cell-free protein synthesis system with comparable protein yields that removes the most expensive substrates and lowers the cell-free reagent cost by over 75% (excluding extract, polymerase, and plasmid) while maintaining high energy levels. This system uses glucose as the energy source and nucleoside monophosphates (NMPs) in place of nucleoside triphosphates (NTPs) as the nucleotide source. High levels of nucleoside triphosphates are generated from the monophosphates within 20 min, and the subsequent energy charge is similar in reactions beginning with either NTPs or NMPs. Furthermore, significant levels (>0.2 mM) of all NTPs are still available at the end of a 3-h incubation, and the total nucleotide pool is stable throughout the reaction. The glucose/NMP reaction was scaled up to milliliter scale using a thin film approach. Significant yields of active protein were observed for two proteins of vastly different size: chloramphenicol acetyl transferase (CAT, 25 kDa) and beta-galactosidase (472 kDa). The glucose/NMP cell-free reaction system dramatically reduces reagent costs while supplying high protein yields.
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PMID:An economical method for cell-free protein synthesis using glucose and nucleoside monophosphates. 1608 Jun 95

A novel strictly aerobic, heterotrophic, pink-pigmented, non-motile, Gram-negative, oxidase-, catalase-, beta-galactosidase- and alkaline phosphatase-positive marine bacterium, designated strain KMM 6058(T), was isolated from the sea urchin Strongylocentrotus intermedius and studied using a polyphasic taxonomic approach. The G+C content of the DNA of the isolate was 41.3 mol%. The predominant fatty acids were i15:1, i15:0, a15:0 and i17:0 3-OH. Phylogenetic analysis based on 16S rRNA gene sequences revealed that strain KMM 6058(T) formed a monophyletic clade with Roseivirga ehrenbergii, with 99% similarity. On the basis of phenotypic, chemotaxonomic, genotypic and phylogenetic characteristics, the novel bacterium should be assigned to the genus Roseivirga as Roseivirga echinicomitans sp. nov. The type strain is KMM 6058(T) (=KCTC 12370(T)=LMG 22587(T)).
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PMID:Roseivirga echinicomitans sp. nov., a novel marine bacterium isolated from the sea urchin Strongylocentrotus intermedius, and emended description of the genus Roseivirga. 1616 68

Sry (sex-determining region on Y chromosome) is expressed in the undifferentiated, bipotential genital ridges of mammalian XY fetuses. The expression of Sry initiates testis development, but the lineage of Sry-expressing cells is unclear. In this study, double-transgenic mice were analyzed using the Cre/loxP system. Cre under the control of the Sry promoter was expressed in the fetal gonads of transgenic mice similarly to endogenous Sry. The Sry/Cre-transgenic mice were crossed with CAG(cytomegalovirus immediate-early enhancer, chicken beta-actin promoter and fusion intron of chicken beta-actin and rabbit beta-globin)/loxP/CAT/loxP/LacZ-transgenic mice, in which the transgene expressed beta-galactosidase after a Cre-mediated recombination event. Sertoli cells, germ cells of testes and granulosa cells of ovaries of double-transgenic mice stained positive with X-gal. Cre expression was detected in germ cells and peritubular/Sertoli cells in adult testes. It is not clear whether beta-galactosidase expression in the Sertoli cells of the testes occurred as a result of Cre expression in the adult or in the fetal gonads. These analyses indicate that cells expressing Sry-inducing factors in female fetal gonads become granulosa cells.
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PMID:Investigation of the fate of Sry-expressing cells using an in vivo Cre/loxP system. 1646 92

Islet neogenesis associated protein (INGAP) promotes the generation of new islet mass in adult animal models. It is not understood what factors control the expression of INGAP. In this study, factors that regulate the expression of INGAP promoter activity are reported. To determine factors that regulate INGAP expression, we previously cloned the promoter region for INGAP. Analysis of the INGAP promoter suggested that candidate regulators of INGAP expression include the transcription factors PDX-1, NeuroD, PAN-1, STAT and AP-1. Using gene addition experiments in the 293 cell line the activity of these transcription factors on an INGAP-promoter construct linked to the beta-galactosidase reporter has been determined. Induction of AP-1 activity or STAT activity using PMA or LIF stimulation respectively, or direct expression of PAN-1 specifically up-regulates INGAP promoter activity. In contrast, co-expression of PDX-1 but not NeuroD inhibits activation of the INGAP-promoter driven by PAN-1, PMA or LIF stimulation. PDX-1 binds directly to the INGAP promoter as determined in electromobility shift and antibody supershift assays. Expression of the INGAP-promoter-reporter construct in the HIT-T15 beta-cell line, a cell line that expresses endogenous PDX-1, did not reveal PMA-mediated stimulation of INGAP promoter activity. HIT-T15 cells however did efficiently transfect (> 68%) and respond (2-fold) to PMA-induced signal transduction to a transfected AP-1-CAT reporter. Partial reduction of PDX-1 expression in HIT-T15 cells was associated with recovery of PMA induced INGAP promoter activity. These data suggest that expression of PDX-1 is associated with a repression of stimulus-induced INGAP promoter activity that appears to be mediated by a direct DNA interaction. These findings implicate PDX-1 in a possible feedback loop to block unbridled islet expansion.
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PMID:PDX-1 can repress stimulus-induced activation of the INGAP promoter. 1652 40


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