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  • 1
    ISSN: 1574-6968
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology
    Notes: Molecular diversity of rumen archaea was analyzed by PCR amplification and sequencing of two 16S rRNA clone libraries prepared from the bovine rumen fluid using two different archaea-specific primer sets. The first library of 19 clones which was generated with primers D30 and D33, produced essentially two groups of sequences, one affiliated with Methanomicrobium mobile (21% of clones) and the other – with the uncultured archaeal sequences from anaerobic digester, which are distantly associated with Thermoplasma (79% of clones). The second library of 25 clones, which was generated with primers 0025e Forward and 1492 Reverse, produced a higher degree of diversity: in addition to the previous two groups, with the M. mobile- (56%) and Thermoplasma-associated sequences (20%), four clones (16%) were identified as Methanobrevibacter spp. The remaining two sequences were associated with unidentified archaeal sequences from the rumen and swine waste. Phylogenetic placement of eight almost complete 16S rRNA sequences revealed the existence of a novel cluster of the rumen Euryarchaeota, which is not affiliated with the known methanogenic archaea.
    Type of Medium: Electronic Resource
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  • 2
    ISSN: 1574-6968
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology
    Notes: The complete nucleotide sequence of a small cryptic plasmid designated pRAO1, from the Gram-negative ruminal bacterium Ruminobacter amylophilus NIAH-3, was determined. The plasmid is a circular DNA molecule, 2140 bp in size, with a GC content of 40%. Computer-assisted analysis identified three open reading frames (ORFs), one of which, ORF3 (347 amino acids), displayed a high degree of amino acid identity with the Mob proteins involved in conjugative mobilization and interplasmid recombination of plasmids from Gram-positive bacteria. We proved the mobilization properties of pRAO1 in the Escherichia coli system using the coresident IncW broad-host-range conjugative plasmid R388. These data demonstrated, for the first time, the mobilization properties of small cryptic plasmids from Gram-negative inhabitants of the rumen.
    Type of Medium: Electronic Resource
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  • 3
    ISSN: 1574-6941
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology
    Notes: Molecular diversity of rumen bacteria was analyzed by PCR amplification and sequencing of 16S rDNA clone libraries prepared from the rumen content of Holstein cows fasted for 16 h. A total of 84 clones, containing almost full size 16S rDNA sequences (about 1.5 kb long), were completely sequenced and subjected to an on line similarity search. Four sequences from the 84 clones closely resembled that of Butyrivibrio fibrisolvens and one clone was found to be related to Treponema bryantii. For 38% of the sequences, the similarity with database sequences was in the range of 90%–98% and for the remaining 56% the similarity was less than 90%. The bacterial community structure was also revealed by phylogenetic placement of sequences in relation to different fractions of rumen content. In the library from the rumen fluid, the sequences were affiliated with the following major phyla: low G+C Gram-positive bacteria (52.4%), Cytophaga-Flexibacter-Bacteroides (38.1%), Proteobacteria (4.7%) and Spirochaetes (2.4%). 2.4% had an uncertain affiliation. The vast majority of sequences from the rumen solids were found to be related to low G+C Gram-positive bacteria (71.4%) and the remaining sequences were placed within the Cytophaga-Flexibacter-Bacteroides (26.2%) and Spirochaetes (2.4%) phyla.
    Type of Medium: Electronic Resource
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  • 4
    ISSN: 1432-0991
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract. Two small cryptic plasmids designated pONE429 and pONE430 were isolated from a rumen bacterium, Selenomonas ruminantium S20. The complete sequence of pONE429 was 2100 bp and contained one open reading frame (ORF) of 201 amino acids. The sequence of pONE430 had 1527 bp and one ORF of 171 amino acids with the similarity of replication protein (Rep protein) of pOM1, pSN2, and pIM13 isolated from Butyrivibrio fibrisolvens, Staphylococcus aureus, and Bacillus subtilis, respectively. In these plasmids, the upstream nucleotide sequence of Rep protein had the conserved nucleotides which could be double-strand origin (DSO) of rolling circle replication (RCR) mechanism. The plasmids of pONE429, pONE430, pJJMI, pJDB21, and pS23 were isolated from S. ruminantium strains and had similar regions that were located within a 〈450-bp nucleotide. These similar regions may be the location that was recognized by the host strain, S. ruminantium.
    Type of Medium: Electronic Resource
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  • 5
    ISSN: 1432-0991
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract A cryptic plasmid designated pSBO2 (3582 bp) was isolated from Streptococcus bovis JB1. The pSBO2 contained putative sites for a double-strand origin (dso), a small transcriptional repressor protein (Cop), countertranscribed RNAs (ctRNAs), and a replication protein (Rep), which were similar to those from pMV158 and pLS1, which were isolated from S. agalactiae, and pWVO1, isolated from Lactococcus lactis. The putative single-strand origin (sso) of pSBO2 was similar to pER341 and pST1, which were isolated from S. thermophilus. Recombinant plasmid designated pSBE2 was constructed to bind pECM184 vector and the DNA fragment containing sso, dso, Cop, ctRNAs, and Rep of pSBO2. When pSBE2 was introduced into S. bovis 12-U-1 and no8, the plasmids in the transformants had deleted the 160-bp fragment between sso and dso. This plasmid, designated pSBE2A, was capable of transforming Escherichia coli and S. bovis strains 12-U-1 and no8 on high frequency; therefore, pSBE2A is an effective shuttle vector.
    Type of Medium: Electronic Resource
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  • 6
    ISSN: 1432-0991
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract Four ruminal Prevotella type strains, P. ruminicola JCM8958T, P. bryantii B14T, P. albensis M384T, and P. brevis ATCC19188T, were characterized for polysaccharide-degrading activities with the reducing sugar release assay and zymogram analyses. Carboxymethylcellulase, xylanase, and polygalacturonate (PG)-degrading enzyme activities were determined in cultures grown on oat spelt xylan, xylose, arabinose, cellobiose, and glucose as sole growth substrates. P. ruminicola and P. albensis showed carboxymethylcellulase induction patterns. When xylan was supplied as a sole growth substrate, xylanase activities produced by P. bryantii and P. albensis were at least 18- and 11-fold higher, respectively, than during growth on other carbohydrates, suggesting that the regulation of the xylanases was highly specific to xylan. All strains constitutively produced PG-degrading enzymes. The corresponding activity of P. bryantii was more than 40-fold higher than in other strains. Zymogram analyses routinely detected the presence of high-molecular-weight (100–170 kDa) polysaccharide-degrading enzymes in ruminal Prevotella. Characteristics of the polysaccharide-degrading activities showed diversity of ruminal Prevotella species.
    Type of Medium: Electronic Resource
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