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:3.1.30.2 (endonuclease)
18,621 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

DNA fingerprinting of 15 reference strains and 24 clinical isolates of Chlamydia trachomatis, 2 strains of C. psittaci and one strain of C. pneumoniae was studied by use of universal 16 + 23S RNA from Escherichia coli, 16S rDNA-directed oligonucleotide and randomly cloned chlamydial DNA probes. The rRNA-gene restriction patterns (ribotypes) enabled the differentiation of chlamydial species. Following DNA cleavage by restriction endonuclease PvuII, lymphogranuloma venereum and trachoma biovars of C. trachomatis could be differentiated. An oligonucleotide, designed to hybridize the C. trachomatis 16S rDNA, also allowed for both species-specific identification and biovar typing of C. trachomatis human strains. Molecular typing system using 3 lambda clones containing C. trachomatis serotype E random DNA inserts, combined to ribotyping, revealed 12 groups of variable banding patterns within C. trachomatis, and could provide an alternative epidemiological tool.
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PMID:DNA fingerprinting of Chlamydia trachomatis by use of ribosomal RNA, oligonucleotide and randomly cloned DNA probes. 129 28

Specific and sensitive amplification of major outer membrane protein (MOMP) gene (ompA) DNA sequences of Chlamydia species with various MOMP genotypes was achieved by a two-step polymerase chain reaction (PCR). Degenerate, inosine-containing oligonucleotide primers homologous to the 5' and 3' ends of the translated regions of all chlamydial MOMP genes were used in a PCR to amplify a DNA fragment of approximately 1,120 bp. A portion of this DNA fragment was amplified in a second genus-specific reaction that yielded a DNA fragment of approximately 930 bp. A pair of degenerate oligonucleotide primers homologous to internal sequences of the primary DNA fragment was used in this PCR. This method detected three cognate chlamydial genomes in a background of 1 microgram of unrelated DNA. MOMP genes of 13 representative chlamydial MOMP genotypes of the species C. trachomatis, C. pneumoniae, and C. psittaci were amplified. In a secondary PCR, group-specific detection was achieved by the simultaneous use of one genus-specific primer and three primers derived from different fingerprint regions of three major groups of chlamydiae. This multiplex PCR differentiated the groups by the length of the amplified DNA fragments and detected the simultaneous presence of DNA sequences of the Chlamydia spp. with different MOMP genotypes. Further differentiation as ompA restriction fragment length polymorphism types among all chlamydial strains with the various MOMP genotypes analyzed here was achieved by restriction endonuclease analysis of the secondary PCR products. DNA sequences corresponding to the ompA restriction fragment length polymorphism type B577 of C. psittaci were detected in two of seven milk samples from cases of bovine mastitis.
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PMID:Two-step polymerase chain reactions and restriction endonuclease analyses detect and differentiate ompA DNA of Chlamydia spp. 134 99

Eight strains of Chlamydia psittaci isolated from swine with pneumonia, pleuritis, pericarditis, and enteritis were characterized through analysis of the major outer membrane protein gene ompA by a two-step polymerase chain reaction, by their interactions with cells in culture, and by the morphologic features and ultrastructure of intracellular inclusions. Amplified chlamydial ompA DNA fragments were differentiated by restriction endonuclease digestion. Chlamydial isolates were separated into 2 types on the basis of ompA restriction fragment length polymorphism. Strains of type L71 had finely granular inclusions, whereas those of type 1710S contained pleomorphic reticulate bodies (RB) in the inclusions, which are characteristic of aberrant chlamydial developmental forms. Chlamydial types L71 and 1710S required centrifuge-assisted inoculation for efficient infection of cell cultures. Cultivation in cell culture medium containing cycloheximide increased the numbers of chlamydial inclusions about 1.5-fold. These strains formed few elementary bodies in yolk sac cells of chicken embryos. Ultrastructurally, unique doublet RB were observed, particularly in strains of the ompA type L71. These doublets consisted of 2 RB, bounded by a cytoplasmic membrane, contained within a common cell wall and an extended periplasmic space. Ultrastructural examination of strains of the ompA type 1710S confirmed the aberrant chlamydial developmental forms, but evidence of viral infection of the RB as a cause of these aberrant forms was not found. The strain S45 isolated from intestinal sites of swine was a trachoma restriction fragment length polymorphism type. With the mouse biotype, it represented the second isolate from animals of Chlamydia trachomatis.
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PMID:Biological properties and genetic analysis of the ompA locus in chlamydiae isolated from swine. 135 14

We constructed the physical map of Chlamydia trachomatis serovar L2 by using three restriction endonucleases, NotI (GC[GGCCGC), SgrAI (C(A/G)[CCGG(T/G)G), and Sse8387I (CCTGCA[GG), and we analyzed the fragments by pulsed-field gel electrophoresis. A total of 25 restriction endonuclease sites and 13 genes and/or operons were located on the map. The genome size was determined to be 1,045 kb. Neither highly transcribed chlamydia genes nor developmental cycle-specific genes were clustered on the genome.
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PMID:Construction of physical and genetic maps of Chlamydia trachomatis serovar L2 by pulsed-field gel electrophoresis. 156 6

DNA from 20 pathogenic or non-pathogenic ruminant strains of Chlamydia psittaci was compared by restriction endonuclease analysis. The strains could be easily differentiated according to their invasiveness for mouse, whatever their pathological origin. DNA patterns of invasive strains were similar, whereas those of non-invasive strains were distributed in two groups.
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PMID:Restriction endonuclease analysis of DNA from ruminant Chlamydia psittaci and its relation to mouse virulence. 162 75

A procedure was developed for characterization of Chlamydia trachomatis strains by using restriction endonuclease analysis of amplified genes of the major outer membrane protein (MOMP). Reference strains of the 15 serovars (A through K and L1 through L3) and clinical isolates were tested. The nucleotide sequences of the MOMP genes of each of the 15 serovars were arbitrarily constructed by using the sequences of the four variable domains known for each serovar and the constant domains of serovar L1. Computer analysis of these sequences indicated that two restriction digestions performed in parallel, one with AluI and the other with IIpaII, followed by HinfI and EcoRI, would allow the theoretical differentiation of 13 serovars. Serovars Ba and L1 presented the same theoretical restriction profile. Our typing method consisted of polymerase chain reaction amplification of a fragment of about 1,200 bp of the MOMP gene, followed by restriction endonuclease digestion with the aforementioned enzymes. From the 15 serovars, we obtained 14 different patterns; 13 profiles were serovar specific, while serovars B and Ba presented the same pattern. Application of this typing method to C. trachomatis strains isolated from clinical material gave the same results as the immunotyping method for 14 of 17 strains. Furthermore, restriction endonuclease analysis detected differences within a serovar. This method seems to be promising for epidemiological studies.
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PMID:Typing of Chlamydia trachomatis by restriction endonuclease analysis of the amplified major outer membrane protein gene. 167 40

A lambda gt11 recombinant library of Chlamydia trachomatis serovar L2 chromosomal DNA was screened with a 29-mer synthetic oligonucleotide specific to the N-terminal amino acids of a predominant 18-kDa chlamydial protein. One recombinant clone, designated lambda gt11/L2/RKA10, was selected on the basis of its strong hybridization signal. Restriction endonuclease analysis and complete nucleotide sequencing of the recombinant revealed a 2,633-bp insert containing one complete open reading frame (ORF2) and two partial ORFs (ORF1 and ORF3). The deduced amino acid sequence of ORF2 matched perfectly at its N-terminal end with the derived amino acid sequence. The 375-bp ORF is capable of encoding a protein comprising 125 amino acids with a molecular mass of 13,689. A sequence compatible with a Shine-Dalgarno ribosome-binding site was located 9 bp upstream from the initiation codon, while the sequence distal to ORF2 revealed a rho-independent terminator. The protein, designated CTH1, possesses an estimated pI of 10.71 due to its high lysine content. This highly basic protein contains no tryptophan or phenylalanine. A protein data base search identified significant homology between CTH1 and painted sea urchin histone H1. Northern (RNA) blot analysis of Chlamydia-infected host cells demonstrated transcripts at 12 h postinfection. The recombinant plasmid encoding ORF2 expressed a gene product of approximately 18 kDa, similar to the native chlamydial protein as analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. This protein appears to represent one of the few eukaryotic histonelike proteins described to date in prokaryotes.
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PMID:Identification and nucleotide sequence of a developmentally regulated gene encoding a eukaryotic histone H1-like protein from Chlamydia trachomatis. 170 78

Both clinical and epidemiological data suggest that inapparent infection by Chlamydia trachomatis occurs in humans. To confirm and study such infections, we developed a hybridization screening system directed toward chlamydial ribosomal RNA (rRNA). Six restriction endonuclease fragments derived from the cloned rrnA operon of chlamydial serovar L2(434) were tested as hybridization screening probes, but only one fragment encoding the 5' portion of the 16s rRNA gene plus some upstream flanking sequence was both sensitive and highly specific in such experiments. In Northern slot blot assays, hybridization analyses with this fragment as probe routinely detected one picogram or less of chlamydial RNA when that RNA was bound to membranes alone or as part of a mixture with a vast excess of mammalian RNA. The probe did not hybridize to RNA from mammalian and relevant bacterial sources but did hybridize to rRNA from B (ocular) and E (genital) serovars of C. trachomatis. Experiments using RNA from conjunctival biopsies and standard conjunctival swab samples from cynomolgus monkeys showed that the probe reliably distinguishes between known chlamydia-infected and uninfected samples. This suggests that it may be useful for clinical screening. Characterization assays for the RNA-directed probe screening system in this monkey model of trachoma provide initial molecular evidence that ocular chlamydial infection may persist longer than previously thought, based solely on direct fluorescence antibody assay (DFA) and culture analyses.
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PMID:RNA-directed molecular hybridization screening: evidence for inapparent chlamydial infection. 175 Apr 44

Specific and sensitive amplification of major outer membrane protein (MOMP) gene DNA sequences of Chlamydia psittaci was achieved in a two-step polymerase chain reaction. First, oligonucleotide primers specific for 5' and 3' nontranslated regulatory regions of the MOMP gene were used in a polymerase chain reaction to amplify a DNA fragment of approximately 1,400 bp. A portion of this DNA fragment was amplified in a second reaction using a degenerate oligonucleotide primer specific for a DNA sequence contained within the 1,400-bp DNA fragment and one of the first-step primers. This method detected 10 cognate chlamydial genomes. C. psittaci MOMP genes from two avian strains and from mammalian serovars 1, 7, and 8 were amplified and analyzed by restriction endonuclease digestion. MOMP genes from mammalian serovars 2 through 6 and 9 and from strains of C. trachomatis and C. pneumoniae could not be amplified. Restriction endonuclease analysis with HaeIII indicated a close relationship between C. psittaci strains of avian and mammalian serovar 1 lineage, while those of mammalian serovars 7 and 8 exhibited distinct restriction patterns. DNA sequences corresponding to the mammalian serovar 1-wild type parakeet MOMP genotype of C. psittaci were detected in two of seven milk samples from cases of bovine mastitis.
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PMID:Detection and strain differentiation of Chlamydia psittaci mediated by a two-step polymerase chain reaction. 177 23

Avian Chlamydia psittaci isolates were examined by restriction endonuclease analysis and serovar-specific monoclonal antibodies and compared with ovine abortion and polyarthritis isolates. The avian isolates were divided into four serovars (turkey, psittacine, pigeon, and duck) based on their reactivity to the monoclonal antibodies. The DNA digest patterns were similar across the four avian serovars; most bands were identical when the isolates were tested with PstI, BamHI, and EcoRI restriction endonuclease enzymes. The turkey group restriction endonuclease analysis patterns were distinguished from those of the other avian strains by three to four band differences with all enzymes. The duck and pigeon isolates showed only minor DNA pattern differences when compared with the psittacine isolates. Four psittacine isolates from various locations in Texas had an extra band with the EcoRI restriction enzyme, suggesting that they were from a common source; however, they were indistinguishable from the other psittacine isolates when examined with the monoclonal antibodies. The avian isolates were distinctly different from either abortion or polyarthritis isolates by both restriction endonuclease analysis and monoclonal antibody analysis. The data demonstrate that the avian isolates form a distinct group or separate biovar with at least four serovars.
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PMID:Comparison of avian Chlamydia psittaci isolates by restriction endonuclease analysis and serovar-specific monoclonal antibodies. 184 67


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