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At least 217 records · Page 12Linked to original sources
PubMed2001

Nuclear DNA sequence specific to Leishmania (Viannia) subgenus: a molecular marker for species identification.

As shown by RFLP analysis, there is a high variability in the beta-tubulin gene region of Leishmania sp. Such variability has been used in the identification of these parasites, establishing differences between subgenera of New World Leishmania. We have found a region of 500 bp (beta500) upstream of the coding region of the beta-tubulin gene that is present in all strains tested belonging to the L. (Viannia) subgenus. This region apparently is a repetitive sequence and we have shown that it is specific to the Leishmania (Viannia) subgenus. This sequence has no homology with the genomic DNA isolated from either the species belonging to the L. (Leishmania) subgenus or other Kinetoplastida, such as Trypanosoma cruzi, T. brucei, Leptomonas samueli, or Crithidia fasciciulata. The beta500 sequence showed sufficient variation to be used as a molecular marker in the identification of parasites. We established inter- and intrasubgenus differentiation and were able to discriminate at the species level in the Vianna subgenus. A PCR assay confirmed the specificity of the beta500 sequence.

Animals↗
PubMed2005

Molecular and morphological evidence indicates that Pseudorhabdosynochus lantauensis (Monogenea: Diplectanidae) represents two species.

Sequences of the first internal transcribed spacer (ITS-1) and the D1-D3 domains of the large subunit (LSU) of the ribosomal DNA (rDNA) were determined for multiple specimens of 4 operational taxonomic units (OTUs) of the monogenean, Pseudorhabdosynochus lantauensis. OTUs were defined based on their collecting localities, host and/or morphological characteristics. All P. lantauensis specimens of one group (OTUs 1 and 3) differed in their sequences of the ITS-1 and partial LSU rDNA when compared with specimens of a second group (OTUs 2 and 4) by 12% and 2%, respectively. Results of the phylogenetic analyses of the LSU rDNA sequence data showed total (100%) bootstrap support for the separation of P. lantauensis into 2 distinct clades. At least 11 of the 18 nucleotide differences in the LSU sequence between the two P. lantauensis clades were derived (i.e. autapomorphic) characters when the morphologically distinct species, P. epinepheli and P. coioidesis, were used as outgroups. Furthermore, there were several autapomorphic character states for each P. lantauensis clade. This provides sufficient evidence to reject the null hypothesis that P. lantauensis represents a single species. Morphological and morphometric differences between these two clades provided additional strong support for the separation of P. lantauensis into two species. These two parasite species were found to co-exist on one of the two species of serranid fish (i.e. Epinephelus coioides) examined in the South China Sea (Guangdong Province, China).

Animals↗
PubMed2005

ITS1-rDNA-based methodology to identify world-wide hake species of the Genus Merluccius.

Species-specific DNA-based tags are valuable tools for the management of both fisheries and commercial fish products. In this study, we have developed a two-step molecular tool to detect the presence of hake DNA (Merluccius spp.) and to identify the exact hake species present in an blind sample. The first test involves PCR amplification of an ITS1-rDNA fragment of 193 bp using nested primers that are interspecifically conserved in Merluccius spp. and Atlantic cod, Gadus morhua. The second test consists of the PCR amplification of a 602-659 bp DNA fragment spanning part of the ribosomal cluster 18S-ITS1-5.8S and digesting it with four restriction enzymes whose targets map at interspecifically nonconserved sites of the ITS1. Alternatively, the identification of hake species can be achieved by FINS or BLAST, using the nucleotide sequence of either the whole ITS1 sequence or its nested fragment of 193 bp. Because of their high reproducibility and ease of execution, these procedures allow for routine analysis and constitute high reliable tools for the rapid identification of 12 species of hake.

Animals↗
PubMed2000

Taxonomic and phylogenetic analysis of Saprolegniaceae (Oomycetes) inferred from LSU rDNA and ITS sequence comparisons.

The aim of this study was to improve our knowledge about the taxonomy and phylogeny of the family Saprolegniaceae, a group of water molds including several pathogens of plants, fish and crustacea. ITS and LSU rDNA were sequenced for representatives of forty species corresponding to ten genera (Achlya, Aphanomyces, Brevilegnia, Dictyuchus, Leptolegenia, Plectospira, Pythiopsis, Saprolegnia, Thraustotheca). Phenetic and cladistic analyses were then carried out. The species Brevilegnia bispora does not appear to belong to the family Saprolegniaceae. Plectospira myrianda clusters with Aphanomyces spp. and they constitute an ancestral group. (Thraustotheca clavata is closely related to the eccentric species of the genus Achlya. The genus Achlya appears polyphyletic, corroborating more or less the three known subgroups, defined by their sexual spore type (eccentric, centric and subcentric). The achlyoid type of spore dehiscence, shared by Aphanomyces and Achlya genera, is shown to be an ancestral character. The saprolegnioid, dictyoid and thraustothecoid types of spore dehiscence are derived characters but their relative evolutionary positions are not resolved.

Classification↗
PubMed2000

RFLP analysis of the ribosomal internal transcribed spacers and the 5.8S rRNA gene region of the genus Saccharomyces: a fast method for species identification and the differentiation of flor yeasts.

The PCR amplification and subsequent restriction analysis of the region spanning the internal transcribed spacers (ITS1 and ITS2) and the 5.8S rRNA gene was applied to the identification of yeasts belonging to the genus Saccharomyces. This methodology has previously been used for the identification of some species of this genus, but in the present work, this application was extended to the identification of new accepted Saccharomyces species (S. kunashirensis, S. martiniae, S. rosinii, S. spencerorum, and S. transvaalensis), as well as to the differentiation of an interesting group of Saccharomyces cerevisiae strains, known as flor yeasts, which are responsible for ageing sherry wine. Among the species of the Saccharomyces sensu lato complex, the high diversity observed, either in the length of the amplified region (ranged between 700 and 875 bp) or in their restriction patterns allows the unequivocal identification of these species. With respect to the four sibling species of the Saccharomyces sensu stricto complex, only two of them, S. bayanus and S. pastorianus, cannot be differentiated according to their restriction patterns, which is in accordance with the hybrid origin (S. bayanus x S. cerevisiae) of S. pastorianus. The flor S. cerevisiae strains exhibited restriction patterns different from those typical of the species S. cerevisiae. These differences can easily be used to differentiate this interesting group of strains. We demonstrate that the specific patterns exhibited by flor yeasts are due to the presence of a 24-bp deletion located in the ITS 1 region and that this could have originated as a consequence of a slipped-strand mispairing during replication or be due to an unequal crossing-over. A subsequent restriction analysis of this region from more than 150 flor strains indicated that this deletion is fixed in flor yeast populations.

Cluster Analysis↗
PubMed2001

Molecular taxonomy and biodiversity of rock fungal communities in an urban environment (Vienna, Austria).

The diversity of fungal communities on three different historical monuments in the city of Vienna (Austria) was analyzed and compared to the fungal diversity of microfungi on rock in the original quarry located in a rural area (Zogelsdorf, Austria). The fungal strains isolated were characterized by morphology and the complete rock fungal community was identified based on molecular data, that is, by sequencing parts of the small ribosomal subunit (18S) and internal transcribed spacer region 1 (ITS1). The genera Coniothyrium, Epicoccum and Phoma were found to be dominant on monument and rock surfaces. Additionally, black yeasts such as Exophiala species and microcolonial fungi like Sarcinomyces and Coniosporium which hitherto were regarded as typical rock inhabitants in semi-arid environments are frequently found on all rock surfaces in Vienna. The biodiversity of the fungi in the urban environment was much higher than on the same rock type in a rural environment, this difference can be attributed to the elevated organic pollution in the city.

Austria↗
PubMed2003

Molecular relatedness within the "Ascochyta pinodes-complex".

The morphological, molecular and pathological characters of 34 isolates identified as Ascochyta pinodes, A. phaseolorum, and Phoma medicaginis var. pinodella were determined. Initial molecular findings, based on PCR fragment size and RFLPs of sections of the mitochondrial genome, the rRNA gene cluster and the beta-tubulin gene complex, were further investigated by sequencing of the total rRNA ITS regions. Isolates were homogenous for all characteristics determined, with the exception of conidial size and septation, and RFLPs of AT rich (presumptive mitochondrial) DNA. The results suggest that the isolates represent host preferential forms of a single taxon, and this finding is discussed in relation to the known teleomorphs and reference material of the species studied.

Ascomycota↗
PubMed2004

Description of two new species of Glypthelmins Stafford, 1905 (Digenea: Macroderoididae) in Rana spp. from Mexico, based on morphology and mtDNA and rDNA sequences.

Glypthelmins Stafford, 1905 includes 29 putative species commonly found in the intestine and liver of anurans from all over the world but mainly in the Americas. Partial sequences of the cytochrome c oxidase subunit 1 ( cox 1), ribosomal internal transcribed spacer region 2 (ITS2) and the large subunit 28S rDNA gene were obtained and analysed using pairwise distance matrices and parsimony methods in order to characterise the interrelationships between 14 isolates of four nominal species of Glypthelmins recognised on morphological grounds. The highest intra-specific sequence divergence occurred in the cox 1 (18.53%) sequence, followed by that of the ITS2 (5.44%) and 28S (4.63%). Genetic variability was detected between the three isolates originally identified as G. facioi Brenes et al., 1959 from two localities in Mexico and one locality in Costa Rica. Sequence divergence exhibited among these isolates ranged from 10.70 to 11.22%, from 0.48 to 0.97% and from 1.33 to 1.88% for cox 1, ITS2 and 28S, respectively. Phylogenetic analysis combining all three data-sets generated a single most parsimonious tree. The three isolates of G. facioi form a clade, with an isolate collected from frogs in Veracruz State as the sister group to an isolate from Tabasco State + G. facioi from Costa Rica. The information derived from pairwise distance of independent data-sets plus the phylogenetic information indicate that each of the two isolates from Mexico, identified a priori as G. facioi, represent separate species. A re-examination of specimens was carried out and a re-evaluation made of the morphological characters to find reliable differences that had been overlooked. As a consequence, G. brownorumae n. sp. from Tabasco and G. tuxtlasensis n. sp. from Veracruz are described based on molecular and morphological differences.

Animals↗
PubMed2006

Characterization of the Drosophila melanogaster genome at the nuclear lamina.

The nuclear lamina binds chromatin in vitro and is thought to function in its organization, but genes that interact with it are unknown. Using an in vivo approach, we identified approximately 500 Drosophila melanogaster genes that interact with B-type lamin (Lam). These genes are transcriptionally silent and late replicating, lack active histone marks and are widely spaced. These factors collectively predict lamin binding behavior, indicating that the nuclear lamina integrates variant and invariant chromatin features. Consistently, proximity of genomic regions to the nuclear lamina is partly conserved between cell types, and induction of gene expression or active histone marks reduces Lam binding. Lam target genes cluster in the genome, and these clusters are coordinately expressed during development. This genome-wide analysis gives clear insight into the nature and dynamic behavior of the genome at the nuclear lamina, and implies that intergenic DNA functions in the global organization of chromatin in the nucleus.

Animals↗
PubMed2002

Evolution and structure of 5S rDNA loci in allotetraploid Nicotiana tabacum and its putative parental species.

Nicotiana tabacum (tobacco) is an allotetraploid derived from ancestors of the modern diploids, N. sylvestris and N. tomentosiformis. We identified and characterized two distinct families of 5S ribosomal DNA (rDNA) in N. tabacum; one family had an average 431 bp unit length and the other a 646 bp unit length. In the diploid species, N. sylvestris and N. tomentosiformis, the 5S rDNA unit lengths are 431 bp and 644 bp respectively. The non-coding spacer sequence of the short unit in tobacco had high sequence homology to the spacer of N. sylvestris5S rDNA, while the longer spacer of tobacco had high homology with the 5S spacer of N. tomentosiformis. This suggests that the two 5S families in tobacco have their origin in the diploid ancestors. The longer spacer sequence had a GC rich sub-region (called the T-genome sub-region) that was absent in the short spacer. Pulsed field gel analysis and fluorescent in situ hybridization to tobacco metaphase chromosomes showed that the two families of 5S rDNA units are spatially separate at two chromosomal loci, on chromosomes S8 (short family) and T8 (long family). The repeat copy number at each chromosomal locus showed heterogeneity between different tobacco cultivars, with a tendency for a decrease in the copy number of one family to be compensated by an increase in the copy number of the second family. Sequence analysis reveals there is as much diversity in 5S family units within the diploid species as there is within the T and S-genome 5S family units respectively, suggesting 5S diversification within each family had occurred before tobacco speciation. There is no evidence of interlocus homogenization of the two 5S families in tobacco. This is therefore substantially different to 18-26S rDNA where interlocus gene conversion has substantially influenced most sequences of S and T genome origin; possible reasons are discussed.

DNA, Intergenic↗
PubMed2001

The internal transcribed spacers (ITS1 and ITS2) of the rDNA differentiates the bark beetle forest pests Tomicus destruens and T. piniperda.

The bark beetles species Tomicus destruens and T. piniperda constitute one of the main pests of European and Asian forests. T. destruens has been thought to be the same species as T. piniperda by some authors. A rapid PCR-based method using internal transcribed spacers (ITS1 and ITS2) of the ribosomal DNA has been developed in order to identifying these two species. We have studied differences in the size of the ITS1, and differences in the sequence and restriction patterns of the ITS2 in 75 individuals of both species. The results indicate a size difference of about 100 bp in the ITS1 of T. destruens and T. piniperda. The size of the ITS2 spacer is similar in both species (approximately 600 bp). However, the sequence is consistently different between T. destruens and T. piniperda in all populations analysed, showing some characteristic indels. Differences in the restriction target in both species for the enzymes HincII and DraI produce different band patterns in agarose NuSieve 5% gel electrophoresis. These data suggest the validity of T. destruens as a phylogenetically differentiated entity from T. piniperda. The phylogenetic analysis of a 789 bp fragment of the 3' end of the cytochrome oxidase I gene (COI) agrees with the two groupings obtained with the ITS2, showing a phylogenetic clustering rather than a phylogeographic structure.

Animals↗
PubMed2002

Population genetic structure of Tomicus piniperda L. (Curculionidae: Scolytinae) on different pine species and validation of T. destruens (Woll.).

Genetic diversity and population structure of Tomicus piniperda was assessed using mitochondrial sequences on 16 populations sampled on 6 pine species in France. Amplifications of Internal transcribed space 1 (ITS1) were also performed. Our goals were to determine the taxonomic status of the Mediterranean ecotype T. piniperda destruens, and to test for host plant or geographical isolation effect on population genetic structure. We showed that T. piniperda clusters in two mtDNA haplotypic groups. Clade A corresponds to insects sampled in continental France on Pinus sylvestris, P. pinaster and P. uncinata, whereas clade B gathers the individuals sampled in Corsica on P. pinaster and P. radiata and in continental France on P. pinea and P. halepensis. Insects belonging to clade A and clade B also consistently differ in the length of ITS1. Individuals belonging to both clades were found once in sympatry on P. pinaster. Genetic distances between clades are similar to those measured between distinct species of Tomicus. We concluded that clade B actually corresponds to the destruens ecotype and forms a good species, T. destruens. Analyses of molecular variance ( amova ) were conducted separately on T. destruens and T. piniperda to test for an effect of either geographical isolation or host species. Interestingly, the effect of host plant was significant for T. piniperda only, while the effect of geographical isolation was not. Pine species therefore seems to act as a significant barrier to gene flow, even if host race formation is not observed. These results still need to be confirmed by nuclear markers.

Animals↗
PubMed2001

Intraindividual and intraspecies variability of ITS1 sequences in the ancient asexual Darwinula stevensoni (Crustacea: Ostracoda).

The lack of variability in ITS regions within individuals and within species has been explained as the result of concerted evolution. In fact, many examples of intraindividual variation in the ITS regions have been reported. Here we report evidence of within-individual variation of the ITS1 region in the obligate parthenogenetic species Darwinula stevensoni. We analysed 46 clones obtained from 12 individuals of D. stevensoni, from three Italian sites and one site in Luxembourg. Seven nucleotides out of 366 were variable. Most variability (80%) was found among clones within individuals, and the remainder of the variability was observed among individuals. No difference was found among populations or between habitats. The low intraspecific variability and the observation of recombinant molecules are evaluated in light of the relevant literature. The high percentage of variation within individuals and the occurrence of recombination without meiosis are discussed by considering the ancient asexual "status" of the species.

Analysis of Variance↗
PubMed1999

High genetic diversity in a population of the ectomycorrhizal basidiomycete laccaria amethystina in a 150-year-old beech forest

The genetic structure of a population of the ectomycorrhizal basidiomycete Laccaria amethystina (Bolt. ex Hooker) Murr. was assessed in a closed 150-year-old beech (Fagus sylvatica L.) forest in the Vosges Mountains in northeastern France. During the autumn of 1994 and 1997, sporophores were collected from three 100-m2 sampling plots located along a 120-m transect crossing the beech stand. The genetic variation of 676 sporophores was initially estimated using heteroduplex analysis of the ribosomal DNA intergenic spacer (IGS1). Ten unique IGS1 heteroduplex/homoduplex patterns were identified, although three types represented most of the sporophores analysed. Each group of IGS1 type was then analysed using random amplified microsatellite analysis (RAMS). RAMS resolved 388 different genotypes amongst the 634 sporophores analysed from the three plots during the autumn of 1994 and 1997. Density as high as 130 genets per 100 m2 was observed during the autumn of 1994. The largest clone covered approximately 1 m2, but most genets covered a few cm2 and produced only one to three sporophores. Only eight genotypes identified in 1994 were found in 1997. Although L. amethystina has the capacity for vegetative persistence, the present study indicates that its populations maintain a genetic structure more consistent with a high frequency of sexual reproduction. This suggests that beech trees could be recolonized by new genotypes each year. Alternatively, this spatial distribution may also arise from erratic fruiting of underground persistent genets. These features (i.e. numerous genets of small size), typical of ruderal species, contrast with studies carried out on other ectomycorrhizal basidiomycetes occurring in mature closed forests.

Journal Article↗
PubMed2002

Detection of genetically divergent clone mates in apomictic dandelions.

This study aims to identify genetically diverged clone mates in apomictic dandelions. Clone mates are defined as individuals that may have diverged as a result of mutation accumulation and that have undergone only clonal reproduction since their most recent common ancestor. Based on distinctive morphology and an aberrant and rare chloroplast haplotype, northwest European individuals of Taraxacum section Naevosa are well suited for the detection of clonal lineages in which mutation has occurred. In the case of strictly clonal reproduction, nuclear genetic variability was expected to be hierarchically organized. Nucleotide polymorphisms in internal transcribed spacer (ITS) sequences, however, were incompatible with a clonal structure of the Norwegian individuals, probably due to persistent ancestral polymorphisms that pre-date the origin of the Naevosa clone. This interpretation is supported by the presence of ITS variants in section Naevosa that were also found in distantly related dandelions. In contrast to the ITS sequence data, amplified fragment length polymorphisms (AFLPs), isozymes and microsatellites strongly supported the contention of prolonged clonal reproduction and mutation accumulation in Norwegian Naevosa. Because these markers are generally considered to be more variable and more rapidly evolving than ITS sequences, mutations in these markers probably evolved after the origin of the clone. Within the Norwegian clone, a surprising number of markers distinguished the clone mates. As a consequence, incorporation of mutation in the detection of clone mates is anticipated to have a big impact on estimates of size, geographical range and age of clones as well as on experimental designs of studies of clonal plants.

Asteraceae↗
PubMed2002

Internal transcribed spacer repeat-specific primers and the analysis of hybridization in the Glycine tomentella (Leguminosae) polyploid complex.

Polyploid and diploid hybridization is a ubiquitous and evolutionarily important phenomenon in the plant world. Determining the parental species of a hybrid, however, is difficult. Molecular markers such as the nuclear ribosomal DNA gene complex, particularly its internal transcribed spacer (ITS) region, have proved powerful in determining hybrid parentage. In some cases, population and genomic phenomena, such as genetic drift and concerted evolution, result in the loss of all or many of the tandemly repeated copies derived from one parental species, making the recovery of hybrid history difficult or impossible. Methods such as direct sequencing and cloning are typically used to find ITS sequences contributed from parental species, but are limited in their ability to detect rare repeat types. Here we report that repeat-specific polymerase chain reaction primers can recover rare parental ITS sequences in the Glycine tomentella polyploid complex. In three allopolyploid lineages of this complex, repeat-specific primers reliably detected rare repeats that both direct sequencing and the screening of many cloned sequences failed to detect. Other strategies, such as the use of exclusion primers, may detect rare parental repeat types in hybrids when previous hypotheses regarding the second parental species are lacking.

Base Sequence↗
PubMed2002

Characterization of a bi-directional promoter for divergent transcription of a PHD-zinc finger protein gene and a ran gene in the protozoan pathogen Giardia lamblia.

We showed previously that transcription of the ran gene in Giardia lamblia is regulated by an AT-rich initiator. In the present study, the ran initiator was found to regulate transcription of a neighbouring PHD zinc-finger protein gene. Deletion and scanning mutagenesis of the phd promoter in a firefly luciferase reporter system showed that the promoter activity is determined by multiple single-stranded T-tract DNA elements distributed into a distal domain spanning the ran initiator (-134/-103) and a proximal domain (-88/-48) spanning phd messenger RNA (mRNA) start sites (-74, -55 and -53 relative to the first ATG). The promoter activity is repressed by the single T-tract element on a non-template strand of the ran initiator, and is activated by closely spaced T-tract elements on the opposite strand. The T-tract elements in the phd and ran initiators compete for similar ssDNA binding proteins. Mutation of -47/-42 resulted in dramatic reduction of luciferase activity without changing luciferase mRNA levels, indicating the potential involvement of a regulatory mechanism in PHD protein translation. These findings suggest that G. lamblia uses multiple copies of a T-tract element as both core and distal elements in regulating transcription initiation, and that expression of the phd gene is regulated at multiple levels.

5' Untranslated Regions↗