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Dichelobacter nodosus is the principal causative agent of ovine footrot. Nucleotide (nt) sequences from the D. nodosus genome have been isolated and a series of overlapping λ clones defining vap (virulence-associated protein) regions 1, 2 and 3 have been reported [Katz et al., J. Bacteriol. 176 (1994) 2663-2669]. In the present study, the limits of the virulence-associated (va) DNA around vap regions...
The nucleotide (nt) sequence of an Ustilago maydis (Um) gene that complemented a partially osmotic-remedial temperature-sensitive (or-ts) mutant defective in apical extension growth has been determined. It contained a continuous open reading frame (OFR) predicted to encode a protein of 853 amino acids (aa). The deduced aa sequence was homologous to γ-adaptin, a component of clathrin-coated vesicles...
We have determined the sequence of the Neurospora crassa met-10 + gene and its flanking regions, and have isolated and analyzed cDNA clones for this region. We have identified two closely linked genes transcribed in the same orientation. Themet-10 + gene is the downstream gene; an open reading frame (ORF) derived from five exons encodes a 475-amino-acid protein. The deduced protein...
Mutations in the terminal 8-bp (5'-T 1 G 2 T 3 G 4 G 5 G 6 C 7 G 8 -3') of the inverted repeats of the bacterial transposon, Tn7, were analysed by measuring Tn7 transposition to the attachment site, attTn7. The mutation, C 2 , present at either end of Tn7 reduces transposition only threefold, but in the double mutant, with C...
The cyclin-dependent kinases, most notable of which is cdc2, are key regulators of the cell cycle, and are highly conserved in evolution. We have cloned and analysed three cdc2-related kinase-encoding genes (tbcrk1-3) from the protozoan parasite Trypanosoma brucei. tbcrk1 encodes a 34-kDa protein with 54% amino acid (aa) identity to the human cdc2, tbcrk2 a 39-kDa protein with 49% identity and tbcrk3...
Site-directed mutagenesis of the penDE gene and expression in Escherichia coli has produced recombinant acylcoenzyme A:isopenicillin N acyltransferase (re-AT) containing amino-acid substitutions in the proenzyme cleavage site (↓) region (Asp-Gly 102 ↓ Cys 103 -Thr-Thr). The effect of these substitutions on proenzyme cleavage and AT activity has been investigated...
We constructed and tested a series of cloning vectors designed to facilitate protein production and purification indictyostelium discoideum (Dd). These vectors carry the origin of replication of the Dd high-copy-number plasmid Ddp2, expression cassettes consisting of the strong, constitutive actin (act15) or the inducible discoidin (disIγ) promoters, a translational start codon upstream from a multiple...
We have isolated and characterized genomic and cDNA recombinant plasmids that encode the Dictyostelium discoideum (Dd) ribosomal protein L3 (rpL3). Genomic plasmids were identified using a probe derived from theSaccharomyces cerevisiae TCM1 gene, that encodes the yeast rpL3. The DdL3 gene contains two introns and encodes a protein 398 amino acids in length that shows a high degree of homology to...
The het-e-1 gene of the fungus Podospora anserina is responsible for vegetative incompatibility through specific interactions with different alleles of the unlinked gene, het-c. Coexpression of two incompatible genes triggers a cell death reaction that prevents heterokaryon formation. The het-e1 allele has been cloned to get information on the function of the locus. It encodes a putative 1356-amino-acid...
DNA coding for bacteriophage T7 RNA polymerase (T7-RNAP) was inserted into a positive selection-vector form of the T4 genome, placing it under the control of bacteriophage T4 ipIII promoters. The recombinant T4 T7-RNAP fusion phage retained infectivity and produced T7-RNAP in infected cells. Fusion genes were constructed by insertion into a plasmid containing an iPIII (encoding internal protein...
We report the cloning and characterization of the 23S ribosomal RNA (rRNA)-encoding gene (rDNA) of Bartonella bacilliformis (Bb). The 2821-bp gene is preceded by an 11-nucleotide (nt) inverted repeat (IR) located 81 nt upstream in the tRNA Ala -23S rDNA intergenic spacer. The gene is followed by an 8-nt IR, five nt downstream in the 23S-5S rDNA intergenic spacer. The nt sequence...
The Saccharomyces cerevisiae LYS7 gene has been cloned based on its genetic map position and complementation of a lys7 mutant. The 1453-bp sequence contains an open reading frame (ORF) that predicts a unique 249 amino acid (aa) protein. A Northern blot experiment demonstrated that LYS7 transcription was not regulated by lysine-specific or general aa control mechanisms. To investigate the effects...
To facilitate future genetic studies with Streptococcus pyogenes (Sp), a recA mutant (Rec11) was constructed using a streptococcal integration vector carrying a PCR-derived internal recA fragment. The insertion of the plasmid in the mutant chromosome was identified by Southern hybridization. Resistance to UV and the ability to accept linear DNA transformation by Rec11 were greatly decreased, confirming...
The recF gene from Azotobacter vinelandii (Av) has been cloned by complementation in an Escherichia coli (Ec) recF mutant. The sequence of 1568 bp has been determined and analyzed. It showed an open reading frame of 1092 nt coding for a 364-amino-acid (aa) polypeptide. The comparison of the deduced aa sequence of the recF of Av with those of other bacteria has elicited the presence of the four...
The nuclear gene NAM9 from Saccharomyces cerevisiae (Sc) codes for a protein which, on the basis of sequence homology, was previously postulated to be a mitochondrial (mt) equivalent of the Escherichia coli (Ec) S4 ribosomal protein (r-protein) [Boguta et al., Mol. Cell. Biol. 12 (1992) 402-412]. The mt-r character of the NAM9 product is now confirmed by cross-reaction with the antisera for the Sc...
A set of antibiotic-resistance and promoter-probe cassettes suitable for insertion mutagenesis and generation of transcriptional fusions was constructed. The cassettes contain the aacC1 gene of transposon Tn1696 conferring resistance to gentamicin in a large variety of Gram - and Gram + bacteria. In addition to the antibiotic-resistance gene, a promoterlessEscherichia coli lacZ gene...
The cyaA gene from Bordetella bronchiseptica (Bb), encoding the adenylate cyclase-hemolysin (AC-Hly), has been cloned and its complete nucleotide sequence has been determined. The deduced amino-acid sequence was compared to the AC-Hly from B. pertussis (Bp) and the main differences were found in the C-terminal repeat region of the molecule.
The identification of a xylose reductase (XR)-encoding gene (XYL1) from the xylose-assimilating yeast Kluyveromyces lactis (Kl) is described. XYL1 was isolated as a highly expressed fusion clone from a 'lacZ translational fusion library. DNA sequence analysis revealed an open reading frame (ORF) of 987 bp capable of encoding a polypeptide of 329 amino acids (aa). The deduced aa sequence displays...
We have used the Aspergillus niger (An) aphA gene as a probe and cloned the A. ficuum (Af) SRRC 265 gene encoding an extracellular pH 6.0-optimum acid phosphatase (APase6) from a genomic library. The identity of the Af aphA gene was confirmed and its nucleotide (nt) sequence verified by comparing its deduced amino acid (aa) sequence to that of purified Af APase6. A comparison of the nt sequences...
Site-specific recombination is thought to play an important role in the evolution of multi-resistant plasmids in bacteria, including the human pathogen Staphylococcus aureus (Sa). A mechanism for site- and orientation-specific recombination between large Sa plasmids was identified in Sa strain 1054. A replication-thermosensitive derivative of plasmid pI9789 Tn552 (called pS1) was found to form...
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