Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:2.7.7.6 (RNA polymerase)
 34,946 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

An in vitro transcription initiation system has been developed from nuclei of rapidly growing, non-green tobacco (Nicotiana tabacum) cultured (BY-2) cells. Conditions for nuclear extraction and in vitro transcription reaction have been optimized with a tobacco beta-1,3-glucanase gene, a constitutively expressed gene in BY-2 cells. The in vitro system supports accurate transcription of RNA polymerase II-dependent promoters from not only plant genes (tobacco beta-1,3-glucanase gene, cauliflower mosaic virus 35S promoter) but also animal genes (adenovirus 2 major late promoter, simian virus 40 early major promoter). In addition, this system drives accurate transcription of an RNA polymerase III-dependent Arabidopsis thaliana U6 snRNA gene. As BY-2 cells do not differentiate in response to light or any other stimuli, they would provide a basal transcription system which lacks tissue-specific and light-responsive nuclear signals as well as chloroplast-derived signals. Consequently, the BY-2 cell-free system is unable to transcribe the tomato gene encoding the small subunit of ribulose-1,5-bisphosphate carboxylase/oxygenase (rbcS3C) whose expression is tissue-specific and light-inducible. However, the transcription of rbcS3C was obtained by supplementing the BY-2 system with a nuclear extract of light-grown tomato seedlings. The promoter regions necessary for rbcS transcription was mapped in vitro using a series of 5' deletion mutants. The 351 bp upstream sequence is essential and the further upstream region from -351 to -441 enhances its transcription. The in vitro basal system will be useful to identify specific signals from both the nucleus and chloroplast in green leaves and other organs/tissues.
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PMID:A plant basal in vitro system supporting accurate transcription of both RNA polymerase II- and III-dependent genes: supplement of green leaf component(s) drives accurate transcription of a light-responsive rbcS gene. 788 33

A gene library of the thermophilic eubacterium, Rhodothermus marinus, strain 21, was prepared in pUC18 and used to transform Escherichia coli. Of 5400 transformants, two produced halos on lichenan plates after Congo-red staining. Restriction mapping showed that the two clones shared an overlapping 1200-bp DNA fragment, which was used for DNA sequencing. Five potential methionine (Met) translational-initiation codons were identified. A putative signal peptide of 30 amino acids was identified with a hydrophobic core of nine hydrophobic amino acids. The molecular mass of the mature enzyme was estimated to be 29.7 kDa. A comparison of the primary protein sequence of beta-glucanase of Rhodothermus marinus with other glycosyl hydrolases showed 38.5% identity to the C-terminal part of the beta-1,3-glucanase of Bacillus circulans and limited identity to bacterial endo-beta-1,3-1,4-glucanases. The amino acid sequence showed high similarity to regions surrounding the catalytic Glu residue of bacterial beta-glucanases. A gene fragment of 889 bp containing the catalytic domain was overexpressed in E. coli using the pET23, T7-phage RNA polymerase system. The enzyme showed activity on lichenan, beta-glucan and laminarin but not on CMC cellulose or xylan. The expressed enzyme was purified by heat treatment of the host. The enzyme had a temperature and pH optima of 85 degrees C and pH 7.0, respectively, and was shown to retain full activity after incubation for 16 h at 80 degrees C and have a half life of 3 h at 85 degrees C.
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PMID:Cloning and sequencing of a Rhodothermus marinus gene, bglA, coding for a thermostable beta-glucanase and its expression in Escherichia coli. 792 16

Plant pathogenesis-related proteins, including beta-1,3-glucanses, are believed to be involved in plant defense mechanisms. We have cloned a beta-1,3-glucanse gene from strawberry (Fragaria x ananassa Duch). This gene, designated FaBG2-1, is a Class II glucanase gene composed of two exons and one intron. The location of a 397-base intron in the gene was confirmed by sequencing a partial cDNA clone obtained by using a rapid amplification of cDNA ends procedure. Also, based on the cDNA sequence, the transcription start site of FaBG2-1 was assigned to a -54G residue. Genomic Southern blot analysis indicated that FaBG2-1 is a member of a multi-gene family. Reverse transcriptase-polymerase chain reaction analysis revealed that this beta-1,3-glucanase gene is expressed constitutively in strawberry leaves.
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PMID:Cloning and partial characterization of a beta-1,3-glucanase gene from strawberry. 1501 49