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111.
In this study, four real-time polymerase chain reaction (PCR) primer sets were developed for the 16S rRNA genes of specific ammonia-oxidizing bacteria (AOB) found in activated sludge of sewage treatment systems. The primer sets target two of several sequence types of the Nitrosomonas oligotropha cluster, members within the Nitrosomonas communis cluster, and all members of the Nitrosomonas europaea–Nitrosococcus mobilis cluster. The detection limit of each primer set was in the range of 3×101–6×102 genes reaction−1. Reliable quantification of the target AOB DNA was obtained when the target AOB DNA comprised more than 0.1% of total AOB DNA in the sample. The application of the primer sets to samples taken from five sewage treatment systems showed that, in all systems, the majority of the AOB population was comprised of one sequence type of the N. oligotropha cluster (3.9±1.5×109–1.7±0.5×1010 cell l−1) and, in most systems, followed by members within the N. communis cluster (2.8±0.3×109–1.0±0.1×1010 cell l−1) or/and another sequence type of the N. oligotropha cluster (1.5±0.6×108–5.5±0.5×108 cell l−1). N. europaea–N. mobilis cluster arose solely in small numbers (4.9±0.8×108 cell l−1) in one system. Real-time PCR-amplified products obtained from genomic DNA extracted from samples were verified using clone library, and it revealed that only the target AOB DNA were PCR amplified, without amplification of the nontarget sequences. 相似文献
112.
Efficient integrative transformation of the phytopathogenic fungus Alternaria alternata mediated by the repetitive rDNA sequences 总被引:4,自引:0,他引:4
An attempt was made to transform Alternaria alternata protoplasts using a plasmid vector, pDH25, bearing the Escherichia coli hygromycin B (Hy) phosphotransferase gene (hph) under the control of the Aspergillus nidulans trpC promoter. Transformants arose on a selective medium containing 100 μg Hy/ml. There were two types of transformants, forming large and small colonies on the selective medium. Transformation with one μg of the vector produced an average of 4.5 large colonies and 600 small ones. In large-colony transformants, the vector often integrated into the recipient chromosome in the form of highly rearranged tandem arrays. To increase transformation efficiency, fragments of the highly repetitive ribosomal RNA gene cluster (rDNA) of A. alternata were used to construct four new vectors for homologous recombination system. Use of these vectors gave higher transformation efficiency than the original plasmid. The best vector, pDH25r1a, gave rise to large-colony transformants at a frequency 20 times higher than pDH25. Transformation events in A. alternata with pDH25r1a occured by homologous recombination as a single crossover between the plasmid-borne rDNA segment and its homologue in the chromosome, often giving rise to tandemly repeated vector DNA. 相似文献
113.
Under oxidative stress mediated by H(2)O(2), significant activation of purified aldose reductase from bovine small intestine was observed in the presence of purified thioredoxin from bovine small intestine. 相似文献
114.
Molecular characterization of Bombyx mori cytoplasmic polyhedrosis virus genome segment 4 总被引:6,自引:0,他引:6 下载免费PDF全文
Ikeda K Nagaoka S Winkler S Kotani K Yagi H Nakanishi K Miyajima S Kobayashi J Mori H 《Journal of virology》2001,75(2):988-995
The complete nucleotide sequence of the genome segment 4 (S4) of Bombyx mori cytoplasmic polyhedrosis virus (BmCPV) was determined. The 3,259-nucleotide sequence contains a single long open reading frame which spans nucleotides 14 to 3187 and which is predicted to encode a protein with a molecular mass of about 130 kDa. Western blot analysis showed that S4 encodes BmCPV protein VP3, which is one of the outer components of the BmCPV virion. Sequence analysis of the deduced amino acid sequence of BmCPV VP3 revealed possible sequence homology with proteins from rice ragged stunt virus (RRSV) S2, Nilaparvata lugens reovirus S4, and Fiji disease fijivirus S4. This may suggest that plant reoviruses originated from insect viruses and that RRSV emerged more recently than other plant reoviruses. A chimeric protein consisting of BmCPV VP3 and green fluorescent protein (GFP) was constructed and expressed with BmCPV polyhedrin using a baculovirus expression vector. The VP3-GFP chimera was incorporated into BmCPV polyhedra and released under alkaline conditions. The results indicate that specific interactions occur between BmCPV polyhedrin and VP3 which might facilitate BmCPV virion occlusion into the polyhedra. 相似文献
115.
CNR/Pcdhalpha family proteins are known as synaptic cadherins and Reelin receptors. Here we report the complete genomic sequence and organization of the rat CNR. The rat CNR cluster encodes 15 variable and 3 constant exons. The genomic organizations of the rat, mouse, and human CNR/Pcdhalpha are orthologous. The percentage identity of the coding regions between the rat and the mouse is 93.6% on average at the nucleic acid level, and between rat and human it is 82.8%. The rat CNRs (v1-v13) also contain an RGD motif in the extracellular cadherin 1 domains and cysteine repeats that are characteristic of the transmembrane and cytoplasmic domains of CNR proteins. The number of variable exons in the rat CNR cluster is identical to that of the human. The rat CNR cluster has one more variable exon than is found in laboratory mouse strains, because in the mouse a variable exon located between v7 and v8 is divided by the insertion of a retrotransposon. This exon is not disrupted in the rat, in which it is transcribed. By in silico analysis, CNR/Pcdhalpha was also mapped to rat chromosome 18, but the orientation was opposite for the mouse CNR/Pcdhalpha gene cluster. The relative expression profiles of the rat CNRs (v1-v13) show that all the CNRs are transcribed, but there are variations in the expression ratios among the CNRs. 相似文献
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Shibata N Suto K Ichimura E Yoshimura K Muneo K Tomigami S Morimoto Y Ogata M Yagi T Higuchi Y Yasuoka N 《Protein and peptide letters》2004,11(1):93-96
Hexadecaheme high molecular weight cytochrome c from a sulfate-reducing bacterium, Desulfovibrio vulgaris Miyazaki F has been successfully purified and crystallized. X-ray diffraction data have been collected by the multiple wavelength anomalous dispersion method. The crystal belongs to the space group P2(1)2(1)2(1) with unit-cell parameters a=60.42, b=84.29 and c=144.16 A and contains one molecule per asymmetric unit. 相似文献
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Rao VU Shiraishi H McDermott PJ 《American journal of physiology. Heart and circulatory physiology》2004,286(6):H2195-H2203
Hypertrophic growth of cardiac muscle is dependent on activation of the PKC-epsilon isoform. To define the effectors of PKC-epsilon involved in growth regulation, recombinant adenoviruses were used to overexpress either wild-type PKC-epsilon (PKC-epsilon/WT) or dominant negative PKC-epsilon (PKC-epsilon/DN) in neonatal rat cardiocytes. PKC-epsilon/DN inhibited acute activation of PKC-epsilon produced in response to phorbol ester and reduced ERK1/2 activity as measured by the phosphorylation of p42 and p44 isoforms. The inhibitory effects were specific to PKC-epsilon because PKC-epsilon/DN did not prevent translocation of either PKC-alpha or PKC-delta. Overexpression of PKC-epsilon/DN blunted the acute increase in ERK1/2 phorphorylation induced by the alpha(1)-adrenergic agonist phenylephrine (PE ). Inhibition of PKC-delta with rottlerin potentiated the effects of PE on ERK1/2 phosphorylation. PKC-epsilon/DN adenovirus also blocked cardiocyte growth as measured after 48 h of PE treatment, although the multiplicity of infection was lower than that required to block acute ERK1/2 activation. PE activated p38 mitogen-activated protein kinase as measured by its phosphorylation, but the response was not blocked by PKC inhibitors or by overexpression of PKC-epsilon/DN. Taken together, these studies show that the hypertrophic agonist PE regulates ERK1/2 activity in cardiocytes by a pathway dependent on PKC-epsilon and that PE-induced growth is mediated by PKC-epsilon. 相似文献