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Results for “DNA, Intergenic”

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At least 19 recordsLinked to original sources

PubMed1996

Use of repetitive intergenic DNA sequences to classify pathogenic and disease-suppressive Streptomyces strains.

PCR DNA fingerprinting using repetitive intergenic DNA sequences (rep-PCR) was investigated as a means of differentiating between closely related strains of Streptomyces which were, in some cases, indistinguishable by other classification methods. Our results demonstrated that the majority of strains had unique rep-PCR DNA fingerprints and established that the technique could be a very useful tool in rapidly determining strain identity.

Base Sequence↗

PubMed1998

The nuclear ribosomal DNA intergenic spacers of wild and cultivated soybean have low variation and cryptic subrepeats.

The intergenic ribosomal DNA spacers (IGSs) from cultivated soybean (Glycine max) and wild soybean (Glycine soja) were sequenced and compared with six other legumes. These IGS sequences were 1821 bp in length in G. soja and G. max cultivars Arksoy, Ransom, and Tokyo, and 1823 bp long in the G. max cultivar Columbus. These represent the smallest published plant IGS sequences to data. Two clones from each of the above five cultivars were sequenced and only 22 sites (1.2%) were polymorphic, thereby supporting previous work that showed low genetic variation in cultivated soybean. The amount of variation observed between different clones derived from the same individual was equal to the amount seen between different cultivars. The soybean IGS sequence was aligned with six other published legume sequences and two homologous regions were identified. The first spans positions 706-1017 in the soybean IGS sequence and ends at a putative promoter site that appears conserved among all legumes. The second is located within the 5' external transcribed spacer, spans positions 1251-1823 in soybean, and includes sequences first identified as subrepeats IV-1 and IV-2 in Vicia angustifolia. Sequences homologous to these two subrepeats were identified among all legume species examined and are here designated "cryptic subrepeats" (CS-1, CS-2) given the range in similarity value (79-96% for CS-1 and 60-95% for CS-2). Comparisons of CS-1 and CS-2 sequences within individual species show that divergence (substitutional mutations, insertions, and deletions) is sufficiently high to obscure recognition of the repeat nature of these sequences by routine dot plot analytical methods. The lack of subrepeats in the 5' half of the soybean IGSs raises questions regarding the role they play in transcription termination or enhancement.

Agriculture↗

PubMed2004

MexZ-mediated regulation of mexXY multidrug efflux pump expression in Pseudomonas aeruginosa by binding on the mexZ-mexX intergenic DNA.

MexZ is a transcriptional regulator of the mexXY multidrug transporter operon, which confers aminoglycoside resistance on Pseudomonas aeruginosa. Highly purified MexZ showed direct binding with a specific site of the mexZ-mexX intergenic DNA when probed by a gel retardation assay. Both in vitro chemical cross-linking experiments and an in vivo two-hybrid expression system showed that the active form of MexZ, which is capable of binding the intergenic DNA, appeared to be a dimer. These results explain the mechanism by which MexZ represses transcription of the mexXY operon, but do not explain the substrate-induced hyperproduction of MexXY. The presence of inducer antibiotic in the gel-retardation assay mixture failed to detect altered MexZ-probe DNA interaction suggesting the possible involvement of an additional regulator.

Amino Acid Sequence↗

PubMed2004

Patterns of evolutionary constraints in intronic and intergenic DNA of Drosophila.

We develop methods to infer levels of evolutionary constraints in the genome by comparing rates of nucleotide substitution in noncoding DNA with rates predicted from rates of synonymous site evolution in adjacent genes or other putatively neutrally evolving sites, while accounting for differences in base composition. We apply the methods to estimate levels of constraint in noncoding DNA of Drosophila. In introns, constraint (the estimated fraction of mutations that are selectively eliminated) is absolute at the 5' and 3' splice junction dinucleotides, and averages 72% in base pairs 3-6 at the 5'-end. Constraint at the 5' base pairs 3-6 is significantly lower in the lineage leading to Drosophila melanogaster than in Drosophila simulans, a finding that agrees with other features of genome evolution in Drosophila and indicates that the effect of selection on intron function has been weaker in the melanogaster lineage. Elsewhere in intron sequences, the rate of nucleotide substitution is significantly higher than at synonymous sites. By using intronic sites outside splice control regions as a putative neutrally evolving standard, constraint in the 500 bp of intergenic DNA upstream and downstream regions of protein-coding genes averages approximately 44%. Although the estimated level of constraint in intergenic regions close to genes is only about one-half of that of amino acid sites, selection against single-nucleotide mutations in intergenic DNA makes a substantial contribution to the mutation load in Drosophila.

Animals↗

PubMedDate not supplied

Primary structure of the ribosomal DNA intergenic spacer from the mosquito, Aedes albopictus.

We have determined the primary structure of a 4.7-kb portion of the ribosomal DNA intergenic spacer from cultured cells of the mosquito, Aedes albopictus. Immediately upstream from the 18S rRNA gene was a 753-bp sequence containing two regions similar to known RNA polymerase I promoters, each preceded by potential transcription termination signals. Upstream from this putative promoter region was a 3.15-kb tandem array of 17 direct repeats with a consensus sequence length of 201 bp. The 201-bp repeats contained imperfect antisense duplications of 11-bp core domain regions in the putative RNA polymerase I promoters, and sequences of possible significance in recombination. Farthest upstream of the 18S rRNA gene was an 803-bp region containing two copies each of 34-, 48-, and 64-bp elements separated by apparently unique sequence. This first detailed structural analysis of a ribosomal DNA intergenic spacer from a member of the lower Diptera has revealed features similar to those described for the higher Diptera as well as conserved motifs presumably critical to rRNA transcription.

Aedes↗

PubMed2005

Identification of strongyle eggs from anthelmintic-treated horses using a PCR-ELISA based on intergenic DNA sequences.

The efficacy of five daily fenbendazole (FBZ) treatments was tested against benzimidazole-resistant cyathostomins in naturally infected horses (n=13). Horses were treated with pyrantel embonate (PYR) to remove adult strongyles followed, 7 days later, by a 5-day course of FBZ. The PYR treatment produced an average faecal egg count reduction of 98%. All samples were negative by faecal egg count 7 days after the start of the FBZ treatment. Positive egg counts were observed from 28 days after the start of FBZ treatment and all horses displayed positive faecal egg counts by 77 days after treatment. Strongyle eggs were harvested from the faeces of the horses prior to treatment and then weekly from 42 to 70 days post-treatment. DNA was obtained from eggs in groups of ten. A PCR-ELISA, based on species-specific differences in intergenic DNA sequences, was used to identify the presence of six cyathostomin species. In pre-treatment samples, Cyathostomum catinatum was detected in nine out of the 13 horses and Cylicostephanus longibursatus, Cylicostephanus goldi and Cylicocyclus nassatus, were found in samples from eight animals. Cylicocyclus ashworthi and Cylicocyclus insigne were not detected pre-treatment. After anthelmintic treatment, C. catinatum and C. longibursatus were most frequently detected, followed by C. nassatus, C. goldi and C. ashworthi. C. insigne was detected at only one time point in a sample from a single horse.

Animals↗

PubMed1992

Is higher-order structure conserved in eukaryotic ribosomal DNA intergenic spacers?

Computer-based structural analysis of the ribosomal DNA intergenic spacer (IGS) from the mosquito Aedes albopictus revealed a potential to form strong and extensive secondary structures throughout a 4.7-kilobase (kb) region. The predicted stability of secondary structures was particularly high within a 3.15-kb region containing 17 tandem 201 base-pair subrepeats. Similarly strong secondary structure potential was also found when IGS subrepeats were analyzed from 17 phylogenetically diverse eukaryotes, including vertebrates, invertebrates, and plants. Conservation of higher-order structure potential in the IGS region of ribosomal DNA may reflect evolutionary and functional constraints on chromatin organization, transcriptional regulation of the ribosomal RNA genes, and/or transcript processing and stability.

Aedes↗

PubMed2003

H2A.Z has a function reminiscent of an activator required for preferential binding to intergenic DNA.

H2A.Z has been shown to regulate transcription in yeast, and that function resides in its C-terminal region as the reciprocal portion of H2A cannot substitute for the latter. We show that fusion of a transcriptional activating region to the C-terminal region of H2A, which is substituted for that of H2A.Z, can allow the chimera to fulfil the special role of H2A.Z in positive gene regulation, as well as complement growth deficiencies of htz1delta cells. We further show that the 'transcription' function of H2A.Z is linked to its ability to preferentially localize to certain intergenic DNA regions. Our results suggest that H2A.Z modulates functional interactions with transcription regulatory components, and thus increases its localization to promoters where it helps poise chromatin for gene activation.

DNA, Fungal↗

PubMed1999

Identification of clinically relevant viridans streptococci by analysis of transfer DNA intergenic spacer length polymorphism.

The utility of PCR analysis of transfer DNA intergenic spacer length polymorphism (tDNA-ILP) for the identification to the species level of clinically relevant viridans streptococci was evaluated with a collection of reference strains of 15 species of the salivarius, anginosus, mitis and mutans rRNA homology groups. PCR products generated by using fluorescent, outwardly directed, consensus tDNA primers were analysed by electrophoresis on denaturating polyacrylamide gels and by laser fluorescence scanning. Eleven species showed specific and distinct tDNA patterns: Streptococcus cristatus, Streptococcus gordonii, Streptococcus oralis, Streptococcus mitis, Streptococcus pneumoniae, Streptococcus sanguinis, Streptococcus parasanguinis, Streptococcus anginosus, Streptococcus mutans, Streptococcus criceti and Streptococcus ratti. Indistinguishable patterns were obtained among two groups of species: Streptococcus vestibularis and Streptococcus salivarius on the one hand and Streptococcus constellatus and Streptococcus intermedius on the other. S. mitis strains produced heterogeneous patterns that could be separated into three groups: a group containing S. mitis biovar 1 and two S. mitis biovar 2 groups, one of which clustered with S. parasanguinis strains while the other showed patterns unrelated to other species. These results agree in part with protein electrophoretic analysis showing that S. mitis biovar 2 strains belong to several streptococcal taxa. In conclusion, PCR analysis of tDNA-ILP holds promise for rapid identification of viridans streptococci that are difficult to identify by phenotypic tests.

Bacterial Typing Techniques↗

PubMed1988

Molecular evolution of intergenic DNA in higher primates: pattern of DNA changes, molecular clock, and evolution of repetitive sequences.

A 3.1-kb intergenic DNA fragment located between the psi beta-globin and delta-globin genes in the beta-globin gene cluster was cloned from gorilla, orangutan, rhesus monkey, and spider monkey, and the nucleotide sequence of each fragment was determined. The phylogeny of these four sequences, together with two previously published allelic sequences from humans and one from chimpanzee, was constructed, and the accumulation of mutations in the region was analyzed. The sites of base substitutions are not evenly distributed within the region: two Alu repeats have accumulated 0.21 + 0.02 substitutions/site with 0.15 + 0.008 substitutions/site in the remainder of the fragment. The occurrence of substitutions at neighboring sites is more frequent than would be expected if they were independent. The observed excesses disappear when ancestral -CG- dinucleotide sites are excluded. The phylogenetic relationships of the sequences indicate that the human sequence shares a most recent coancestor with the chimpanzee sequence. The data also show that great apes have accumulated fewer mutations in this part of the genome than has the rhesus monkey. The relative rates of accumulation of 12 kinds of nucleotide substitution in the region during primate evolution are asymmetric in the DNA strands. From these rates of accumulation, the origin of a simple stretch of sequence near the 3' end of the 3.1-kb fragment was deduced to be a sequence comprising 50% T and 50% C on one strand. The two oppositely oriented Alu sequences in the 3.1-kb region were inserted at their present positions before the divergence of the New-World monkeys from other lineages. Our analysis shows that the nucleotide sequences of the two Alu repeats in spider monkey are unexpectedly similar both to each other and to the deduced ancestral sequence of Alu repeats. The data suggest that there has been some type of recombinational event between the spider monkey Alu repeats but that it was not a simple gene conversion.

Animals↗

PubMed1993

Multiple nuclear protein binding to 135 bp subrepeat element of wheat ribosomal DNA intergenic spacer.

Specific binding of wheat nuclear proteins to 135 bp subrepeat element of the ribosomal DNA intergenic spacer was studied by electrophoretic mobility shift assays. A number of specific DNA-protein complexes are readily formed upon incubation of the labelled 135 bp subrepeat element with wheat nuclear protein extracts. Some of these complexes have various DNA-binding specificities as evidenced by their optimal detection in the presence of different non-specific competitor polynucleotides. One of the wheat nuclear proteins was found to bind specifically both to 135 bp subrepeat element and an intergenic spacer region located downstream of the transcription initiation site (between +192 and +366 bp). No apparent effects of the 135 bp element methylation with methylase HpaII on protein binding were found.

Binding, Competitive↗

PubMed2003

Bacillus anthracis diverges from related clades of the Bacillus cereus group in 16S-23S ribosomal DNA intergenic transcribed spacers containing tRNA genes.

Mung bean nuclease treatment of 16S-23S ribosomal DNA intergenic transcribed spacers (ITS) amplified from several strains of the six species of the Bacillus cereus group showed that B. anthracis Davis TE702 and B. mycoides G2 have other intermediate fragments in addition to the 220- and 550-bp homoduplex fragments typical of the B. cereus group. Long and intermediate homoduplex ITS fragments from strains Davis TE702 and G2 and from another 19 strains of the six species were sequenced. Two main types of ITS were found, either with two tRNA genes (tRNA(Ile) and tRNA(Ala)) or without any at all. Strain Davis TE702 harbors an additional ITS with a single tRNA gene, a hybrid between the tRNA(Ile) and tRNA(Ala) genes, suggesting that a recombination event rather than a deletion generated the single tDNA-containing ITS. Strain G2 showed an additional ITS of intermediate length with no tDNA and no similarity to other known sequences. Neighbor-joining analysis of tDNA-containing long ITS indicated that B. cereus and B. thuringiensis represent a single clade. Three signature sequences discriminated B. anthracis from B. cereus and B. thuringiensis, indicating that the anthrax agent started evolving separately from the related clades of the B. cereus group. B. mycoides and B. weienstephanensis were very closely related, while B. pseudomycoides appeared the most distant species.

Bacillus anthracis↗

PubMed1999

Ribosomal DNA intergenic spacer of the swimming crab, Charybdis japonica.

We have determined the full sequence of the ribosomal DNA intergenic spacer (IGS) of the swimming crab, Charybdis japonica, by long PCR for the first time in crustacean decapods. The IGS is 5376 bp long and contains two nonrepetitive regions separated by one long repetitive region, which is composed mainly of four subrepeats (subrepeats I, II, III, and IV). Subrepeat I contains nine copies of a 60-bp repeat unit, in which two similar repeat types (60 bp-a and 60 bp-b) occur alternatively. Subrepeat II consists of nine successive repeat units with a consensus sequence length of 142 bp. Subrepeat III consists of seven copies of another 60-bp repeat unit (60 bp-c) whose sequence is complementary to that of subrepeat I. Immediately downstream of subrepeat III is subrepeat IV, consisting of three copies of a 391-bp repeat unit. Based on comparative analysis among the subrepeats and repeat units, a possible evolutionary process responsible for the formation of the repetitive region is inferred, which involves the duplication of a 60-bp subrepeat unit (60 bp-c) as a prototype.

Animals↗

PubMed2001

Rapid identification of clinically relevant Legionella spp. by analysis of transfer DNA intergenic spacer length polymorphism.

Analysis of PCR-amplified transfer DNA (tDNA) intergenic spacers was evaluated as a rapid method for identification to the species level of 18 species of Legionella known as human pathogens. Type strains (n = 19), reference strains (n = 16), environmental strains (n = 31), and clinical strains (n = 32) were tested. PCR products using outwardly directed tDNA consensus primers were separated on polyacrylamide gels and analyzed with automated laser fluorescence. Test results were obtained in 8 h starting with 72-h-old bacterial growth on solid medium. Species-specific patterns were obtained for all 18 Legionella species tested: Legionella anisa, L. bozemanii serogroups 1 and 2, L. cincinnatiensis, L. dumoffii, L. feeleii serogroups 1 and 2, L. gormanii, L. hackeliae serogroups 1 and 2, L. jordanis, L. lansingensis, L. longbeachae serogroups 1 and 2, L. lytica, L. maceachernii, L. micdadei, L. oakridgensis, L. parisiensis, L. pneumophila serogroups 1 to 14, L. sainthelensi serogroup 2, L. tucsonensis, and L. wadsworthii. Computer-assisted matching of tDNA-intergenic length polymorphism (ILP) patterns identified all 63 environmental and clinical strains to the species level and to serogroup for some strains. tDNA-ILP analysis is proposed as a routinely applicable method which allows rapid identification of environmental and clinical isolates of Legionella spp. associated with legionellosis.

Computational Biology↗

PubMed2005

Simple species identification of Trichinella isolates by amplification and sequencing of the 5S ribosomal DNA intergenic spacer region.

We developed a PCR-based assay using a single primer pair to amplify the 5S ribosomal DNA intergenic spacer region to identify Trichinella isolates. In our method, amplified products are directly sequenced on both strands and compared to GenBank sequences. Using this method, we were able to identify Trichinella spiralis, T. britovi and T. nativa. This method permits rapid species identification of Trichinella isolates; however, further evaluation is required before recommending this approach for routine use.

Animals↗

PubMed2002

The upstream Sal repeat-containing segment of Arabidopsis thaliana ribosomal DNA intergenic region (IGR) enhances the activity of adjacent protein-coding genes.

The sequence containing 'upstream Sal repeats' (USR) from the Arabidopsis thaliana ribosomal DNA intergenic region (IGR) was tested for its influence on the in vivo activity of nearby protein coding genes. On average, the presence of the IGR fragment leads to a four-fold increase in the expression of a reporter gene, beta-glucuronidase, under control of the strong CaMV 35S promoter. With the help of the site-specific cre-lox recombination system, we have also obtained pairs of transgenic lines with or without the USR-containing fragment, both integrated at the same chromosomal position. Results with these transgenic lines, which contain an NPT II (kanamycin resistance) gene under control of the nos promoter as a test gene, confirmed the results obtained with the CaMV 35S-driven GUS gene. Moreover, they show that the IGR sequence can oppose tendencies of gene silencing. We hypothesize that the described effect relates to features of the chromatin structure in the proximity of the upstream Sal repeats.

Arabidopsis↗

PubMed1999

The nuclear ribosomal DNA intergenic spacer as a target sequence to study intraspecific diversity of the ectomycorrhizal basidiomycete Hebeloma cylindrosporum directly on pinus root systems.

Polymorphism of the nuclear ribosomal DNA intergenic spacer (IGS) of the ectomycorrhizal basidiomycete Hebeloma cylindrosporum was studied to evaluate whether this sequence could be used in field studies to estimate the diversity of strains forming mycorrhizas on individual Pinus pinaster root systems. This sequence was amplified by PCR from 125 haploid homokaryotic strains collected in 14 P. pinaster stands along the Atlantic coast of France by using conserved oligonucleotide primers. Restriction enzyme digestion of the amplified 3.4-kbp-long IGS allowed us to characterize 24 alleles whose frequencies differed. Nine of these alleles were found only once, whereas about 60% of the strains contained four of the alleles. Local populations could be almost as diverse as the entire population along a 150-km stretch of coastline that was examined; for example, 13 alleles were found in a single forest stand. The IGS from one strain was partially sequenced, and the sequence data were used to design oligonucleotides which allowed separate PCR amplification of three different segments of the IGS. Most polymorphisms observed among the full-length IGS regions resulted from polymorphisms in an internal ca. 1,500-bp-long sequence characterized by length variations that may have resulted from variable numbers of a T2AG3 motif. This internal polymorphic sequence could not be amplified from the genomes of nine other Hebeloma species. Analysis of this internal sequence amplified from the haploid progenies of 10 fruiting bodies collected in a 70-m2 area resulted in identification of six allelic forms and seven distinct diplotypes out of the 21 possible different combinations. Moreover, optimization of the PCR conditions resulted in amplification of this sequence from more than 80% of the DNA samples extracted from individual H. cylindrosporum infected P. pinaster mycorrhizal root tips, thus demonstrating the usefulness of this sequence for studying the below-ground diversity of mycorrhizas formed by genets belonging to the same fungal species.

Alleles↗

PubMed2005

Ribosomal DNA intergenic spacer sequence in foxtail millet, Setaria italica (L.) P. Beauv. and its characterization and application to typing of foxtail millet landraces.

We determined the sequence of ribosomal DNA (rDNA) intergenic spacer (IGS) of foxtail millet isolated in our previous study, and identified subrepeats in the polymorphic region. We also developed a PCR-based method for identifying rDNA types based on sequence information and assessed 153 accessions of foxtail millet. Results were congruent with our previous works. This study provides new findings regarding the geographical distribution of rDNA variants. This new method facilitates analyses of numerous foxtail millet accessions. It is helpful for typing of foxtail millet germplasms and elucidating the evolution of this millet.

Base Sequence↗