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:2.7.7.49 (reverse transcriptase)
31,746 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Carp acclimated to 10 degrees C gave 69k, 66k, and 62kDa light meromyosin (LMM) fragments in SDS-PAGE, while fish acclimated to 30 degrees C gave 74k, 69k, 66k, and 62kDa fragments. The microsequence analysis revealed that the 69k and 66kDa components from the 10 degrees C-acclimated carp contained an N-terminal amino acid sequence different from that of 62kDa. The four fragments from the 30 degrees C-acclimated carp showed the same sequence as that of the 69k and 66kDa components from the 10 degrees C-acclimated carp, except that the 2nd amino acid, Ala, of the 10 degrees C-acclimated LMM was replaced by Thr. DNA fragments encoding an N-terminal region of LMM were amplified by PCR or reverse transcriptase-PCR, demonstrating that the two acclimated groups further contained several amino acids substituted.
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PMID:Temperature acclimation induces light meromyosin isoforms with different primary structures in carp fast skeletal muscle. 788 20

Total RNA was isolated from carp pituitary gland. The first strand cDNA was synthesized using oligo(dT) 12-18 as a primer, the total RNA as a template, and AMV reverse transcriptase. Next the polymerase chain reaction (PCR) was performed using the first strand cDNA as a template, the synthetic 29 oligonucleotides as primer and Taq DNA polymerase (94 degrees C, 60 s; 55 degrees C, 30 s; 72 degrees C, 50 s; 35 cycles). After PCR amplification, the products were cloned into an E. coli expression vector (PBluescript II KS+/-). The result of the sequence analysis and the restriction map shows that an open reading frame of the carp growth hormone gene contains 630 base pairs which code for a polypeptide of 210 amino acids including 22 amino acids of the signal peptide and 188 amino acids of the nature growth hormone. Nucleotide sequence and amino acid sequence of our carp growth hormone gene are the same as Koren's carp GH cDNA in the coded region. Compared with Chao's carp GH cDNA, the homology of nucleotide sequence and amino acid sequence for our carp growth hormone gene is 95.6% and 96.7%, respectively, in the coded region.
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PMID:Amplification, cloning, and sequence comparison of the growth hormone gene for carp (Cyprinus carpio) by the polymerase chain reaction. 909 58

A cDNA encoding the precursor of one of the major components of gilthead sea bream, Sparus aurata, egg envelope has been cloned by reverse transcriptase polymerase chain reaction (RT-PCR) techniques. The clone was isolated starting from total RNA extracted from the liver of spawning female fish and estradiol-17 beta-treated male fish. Sequence analysis revealed that the cDNA encoded a protein of 405 aa corresponding to 49-kDa component (termed gp49), a glycoprotein belonging to the N-linked type. The gp49 protein is homologous to the Zl-3 of medaka Oryzias latipes, the mammalian ZPC and ZPC homologues of Xenopus laevis (xlZPC) and carp Cyprinus carpio (ccZPC). In addition, the open reading frame also encodes an additional aa sequence, the signal peptide, located in the N-terminal region of the protein. RT-PCR and in situ expression analyses evidenced an organ-restricted pattern: the mRNA was detected only in liver of spawning female and estradiol-17 beta-treated male fish but not in other tissues.
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PMID:Identification and spatial distribution of the mRNA encoding the gp49 component of the gilthead sea bream, Sparus aurata, egg envelope. 940 96

Pro-opiomelanocortin (POMC) is the precursor of a number of biologically active peptides, including adrenocorticotropic hormone, alpha-melanocyte-stimulating hormone and beta-endorphin, which are released by the pituitary glands of fish as well as mammals. To quantify the levels of expression of the two POMC mRNAs relative to one another during the response of the common carp to temperature-induced stress, we used reverse transcriptase PCR combined with capillary electrophoresis and laser-induced fluorescence detection. The ratio of POMC-I mRNA to POMC-II mRNA determined in wild-type and four isogenic carp strains was found to be strain-dependent and influenced by temperature. In strain E20xR8, the ratio had altered in favour of POMC-I from 1:3.2 (POMC-I:POMC-II) in fish adapted to 24 degreesC to 1:1.2 in fish adapted to a decrease of 9 degreesC in ambient temperature. A rapid drop in temperature from 24 to 15 degreesC decreased the POMC mRNA ratio at the expense of POMC-I from 1:1.9 in the control fish (strain E4xR3R8) to 1:4.2 3 h after the temperature drop of 9 degreesC. We conclude that both POMC genes are expressed in the common carp and that their expression ratio is strain-dependent and changes in response to ambient temperature.
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PMID:Differential expression of two pro-opiomelanocortin mRNAs during temperature stress in common carp (Cyprinus carpio L.). 979 45

To determine the ease and feasibility of amplifying the beta-actin gene in fish by the polymerase chain reaction (PCR), genomic DNAs of several fish (Rivulus, Southern top mouth minnow, common fat minnow, oily bitterling, carp, Far Eastern catfish, medaka, and European flounder) were extracted and used as a template with conserved primers, designed on the basis of high amino acid homology (approximately 98% or more). Among them, the self-fertilizing hermaphroditic fish Rivulus marmoratus was chosen for further characterization. After amplification of the Rivulus beta-actin PCR product with Taq polymerase, PCR product was subcloned to pCRII vector. After restriction enzyme mapping of Rivulus beta-actin gene, the amplified insert was sequenced using ALF Express automatic DNA sequencer with conserved internal primers. The R. marmoratus beta-actin gene consists of 1763 bp encoding 375 amino acids including 5 exons and 4 introns. The splicing and acceptance sites of the exon and intron boundaries of the Rivulus beta-actin gene were highly conserved with consensus sequences (GT/AG). The amino acid homology of R. marmoratus beta-actin to other species was high: 98.93% to human; 98.93%, Atlantic salmon; 98.93%, common carp; 98.93%, grass carp; 98.93%, zebrafish; 98.67%, medaka; and 98.40%, sea bream. To determine the expression of the R. marmoratus beta-actin gene in liver and ovary, reverse transcriptase-polymerase chain reaction was carried out with internal primers. In conclusion, these universal primers are successful in the rapid cloning of the fish beta-actin gene by PCR, based on a high homology of the beta-actin gene conserved through evolution. This approach will be applicable to the isolation of other beta-actin homologues in the investigation of phylogenetic comparisons of fish species, along with a possible application to cloning strategy in other conserved genes.
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PMID:The Internally Self-fertilizing Hermaphroditic Teleost Rivulus marmoratus (Cyprinodontiformes, Rivulidae) beta-Actin Gene: Amplification and Sequence Analysis with Conserved Primers. 1081 55

Rex1, together with the related BABAR: elements, represents a new family of non-long-terminal-repeat (non-LTR) retrotransposons from fish, which might be related to the CR1 clade of LINE elements. Rex1/BABAR: retrotransposons encode a reverse transcriptase and an apurinic/apyrimidinic endonuclease, which is very frequently removed by incomplete reverse transcription. Different Rex1 elements show a conserved terminal 3' untranslated region followed by oligonucleotide tandem repeats of variable size and sequence. Phylogenetic analysis revealed that Rex1 retrotransposons were frequently active during fish evolution. They formed multiple ancient lineages, which underwent several independent and recent bursts of retrotransposition and invaded fish genomes with varying success (from <5 to 500 copies per haploid genome). At least three of these ancient Rex1 lineages were detected within the genome of poeciliids. One lineage is absent from some poeciliids but underwent successive rounds of retrotransposition in others, thereby increasing its copy number from <10 to about 200. At least three ancient Rex1 lineages were also detected in the genome project fish Fugu rubripes. Rex1 distribution within one of its major lineages is discontinuous: Rex1 was found in all Acanthopterygii (common ancestor in the main teleost lineage approximately 90 MYA) and in both European and Japanese eels (divergence from the main teleost lineage about 180 MYA) but not in trout, pike, carp, and zebrafish (divergence 100-120 MYA). This might either result from frequent loss or rapid divergence of Rex1 elements specifically in some fish lineages or represent one of the very rare examples of horizontal transfer of non-LTR retrotransposons. This analysis highlights the dynamics and complexity of retrotransposon evolution and the variability of the impact of retrotransposons on vertebrate genomes.
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PMID:Multiple lineages of the non-LTR retrotransposon Rex1 with varying success in invading fish genomes. 1107 55

Estrogen receptor-mediated induction of zona radiata (ZR) and vitellogenin (VTG) mRNA and protein in rainbow trout (Oncorhynchus mykiss) was compared to assess their utility as biomarkers for exposure to estrogenic compounds. Partial sequences of rainbow trout ZR and beta-actin were cloned by reverse transcriptase polymerase chain reaction (RT-PCR) using degenerate primers based on conserved regions across a number of species. A 549 bp fragment of the rainbow trout ZR-gene showed a high degree of amino acid sequence identity to that of salmon (77%), winter flounder (64%), carp ZP2 (63%) and medaka (61%) ZR-proteins. The 1020 bp beta-actin fragment was approximately 100% identical to sequences from several species. Real-time PCR was used to quantify the induction of ZR-gene and VTG in rainbow trout liver after in vivo exposure to estradiol-17 beta (E(2)) (0.01, 0.1, 1.0 or 10 mg/kg body weight (bw) fish) or alpha-zearalenol (alpha-ZEA) (0.1, 1.0 or 10 mg/kg bw). Real-time PCR and indirect enzyme-linked immunosorbent assay (ELISA) showed that ZR and VTG were induced in both the liver and the plasma after a single injection of E(2) or alpha-ZEA. ZR was more responsive to low levels of E(2) and alpha-ZEA than VTG, and real-time PCR was shown to be more sensitive than the ELISA. Rainbow trout ZR-gene and proteins provide a sensitive biomarker for assessing estrogenic activity.
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PMID:Quantification of rainbow trout (Oncorhynchus mykiss) zona radiata and vitellogenin mRNA levels using real-time PCR after in vivo treatment with estradiol-17 beta or alpha-zearalenol. 1122 27

To improve our understanding of the genetic basis of fish disease, we developed a pathogen model, using zebrafish (Danio rerio) and spring virema of carp virus (SVCV). Replicate groups of 10 fish were acclimated to 20 or 24 degrees C, then were exposed to SVCV concentrations of 10(3) to 10(5) plaque-forming units per milliliter (PFU/ml) of water and observed daily. In a second trial, fish were acclimated to 15 degrees C, and replicate groups of 10 fish were exposed to SVCV at a concentration of 10(5) PFU/ml; however, the temperature was raised 1 degrees C/wk. Moribund fish were collected for histologic examination, and dead fish were assayed for virus by use of cell culture and reverse transcriptase-polymerase chain reaction (RT-PCR) analysis. Mortality exceeded 50% in fish exposed to 10(5) PFU of SVCV/ml at the lower temperatures. Clinical signs of disease became evident seven days after viral exposure and were observed most consistently in fish of the 10(5) PFU/ml groups. Affected zebrafish were anorectic and listless, with epidermal petechial hemorrhages followed by death. Use of plaque assays and RT-PCR analysis confirmed presence of SVCV at titers > or = 10(4) PFU/g of tissue. Histologic lesions included multifocal brachial necrosis and melanomacrophage proliferation in gills, liver, and kidneys. These results indicate that zebrafish are susceptible to infection by SVCV under conditions that mimic a natural route of exposure.
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PMID:Susceptibility of zebrafish (Danio rerio) to a model pathogen, spring viremia of carp virus. 1465 94

Rhabdoviruses were isolated from perch Perca fluviatilis and largemouth bass Micropterus salmoides exhibiting clinical signs of disease. Preliminary studies indicated that these viruses could be neutralised by antisera to perch rhabdovirus (Dorson et al. 1984) and may be similar to those previously isolated from grayling Thymallus thymallus and pike-perch Stizostedion stizostedion. The relationship between these viruses and the previously characterised fish rhabdoviruses, pike fry rhabdovirus (PFRV), spring viraemia of carp virus (SVCV) and lake trout rhabdovirus, was investigated. Viruses were propagated in bluegill fry (BF-2) cells and were characterised using electron microscopy, serum neutralisation tests, immunofluorescence tests, sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and nucleotide sequence analysis. The bullet-shaped viral particles appeared to be compact, with spikes visible at the surface, a morphology similar to that of the vesiculovirus group of rhabdoviruses. Serum neutralisation tests showed that the viruses were antigenically closely related to the previously characterised perch rhabdovirus, but were not significantly neutralised by antisera to PFRV, SVCV or viral haemorrhagic septicaemia virus (VHSV). In immunofluorescence tests with perch rhabdovirus antisera, strong specific fluorescence was observed in cell cultures infected with the new rhabdovirus isolates, but no fluorescence was observed with antisera to PFRV, SVCV or VHSV. SDS-PAGE analysis revealed a polypeptide profile typical of vesiculoviruses, but the novel virus isolates had different relative mobilities of their P and M proteins compared to PFRV and SVCV. Nucleotide sequence analysis was carried out using reverse transcriptase-polymerase chain reaction (RT-PCR) and DNA sequencing of a 439 base-pair region of the viral L gene. The novel rhabdovirus isolates had <76% nucleotide sequence identity to PFRV, SVCV and lake trout rhabdovirus and >95% identity to perch rhabdovirus. Phylogenetic analysis using both maximum parsimony and neighbour-joining methods assigned the perch rhaboviruses to a separate group to that of PFRV, SVCV and lake trout rhabdovirus. These data are the initial characterisation of a group of emerging fish vesiculo-type viruses that are biochemically and genetically distinct from the PFRV, SVCV and lake trout rhabdoviruses.
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PMID:Emerging vesiculo-type virus infections of freshwater fishes in Europe. 1496 32

Cytokines and IgM levels are important parameters in the acquired immunity of fish. In the present study, the effect of pentachlorophenol (PCP) on the expression of proinflammatory cytokines, TNF-alpha (tumor necrosis factor-alpha) and IL-1beta (interleukin 1) mRNA levels of crucian carp, were determined by semi-quantitative reverse transcriptase-polymerase chain reaction (RT-PCR) method. To put the deduction of PCP's immunotoxicity on B cell function, B cells secretion of IgM under exposure of PCP-administrated fish macrophage was determined by enzyme-linked immunosorbent assay (ELISA), and the ability of B cells to secrete IgM was determined by enzyme-linked immunospot (ELISPOT). The results showed that the mRNA expressions of these two cytokines were suppressed by the administration of PCP. The supernatants from PCP-administrated fish macrophage showed less stimulation on B cell, lower maturation of B cells and secretion of IgM. These results suggested that PCP might have impact on micromilieu immune factors as proinflammatory cytokines.
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PMID:Pentachlorophenol reduces B lymphocyte function through proinflammatory cytokines in Carassius auratus. 1562 36


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