Family Of Repetitive DNA Sequences Research Articles

Large vs small genomes in Passiflora: the influence of the mobilome and the satellitome

While two lineages of retrotransposons were more abundant in larger Passiflora genomes, the satellitome was more diverse and abundant in the smallest genome analysed. Repetitive sequences are ubiquitous and fast-evolving elements responsible for size variation and large-scale organization of plant genomes. Within Passiflora genus, a tenfold variation in genome size, not attributed to polyploidy, is known. Here, we applied a combined in silico and cytological approach to study the organization and diversification of repetitive elements in three species of this genus representing its known range in genome size variation. Sequences were classified in terms of type and repetitiveness and the most abundant were mapped to chromosomes. We identified long terminal repeat (LTR) retrotransposons as the most abundant elements in the three genomes, showing a considerable variation among species. Satellite DNAs (satDNAs) were less representative, but highly diverse between subgenera. Our results clearly confirm that the largest genome species (Passiflora quadrangularis) presents a higher accumulation of repetitive DNA sequences, specially Angela and Tekay elements, making up most of its genome. Passiflora cincinnata, with intermediate genome and from the same subgenus, showed similarity with P. quadrangularis regarding the families of repetitive DNA sequences, but in different proportions. On the other hand, Passiflora organensis, the smallest genome, from a different subgenus, presented greater diversity and the highest proportion of satDNA. Altogether, our data indicates that while large genomes evolved by an accumulation of retrotransposons, the smallest genome known for the genus has evolved by diversification of different repeat types, particularly satDNAs.

Chromosome size-correlated and chromosome size-uncorrelated homogenization of centromeric repetitive sequences in New World quails

Many families of centromeric repetitive DNA sequences isolated from Struthioniformes, Galliformes, Falconiformes, and Passeriformes are localized primarily to microchromosomes. However, it is unclear whether chromosome size-correlated homogenization is a common characteristic of centromeric repetitive sequences in Aves. New World and Old World quails have the typical avian karyotype comprising chromosomes of two distinct sizes, and C-positive heterochromatin is distributed in centromeric regions of most autosomes and the whole W chromosome. We isolated six types of centromeric repetitive sequences from three New World quail species (Colinus virginianus, CVI; Callipepla californica, CCA; and Callipepla squamata, CSQ; Odontophoridae) and one Old World quail species (Alectoris chukar, ACH; Phasianidae), and characterized the sequences by nucleotide sequencing, chromosome in situ hybridization, and filter hybridization. The 385-bp CVI-MspI, 591-bp CCA-BamHI, 582-bp CSQ-BamHI, and 366-bp ACH-Sau3AI fragments exhibited tandem arrays of the monomer unit, and the 224-bp CVI-HaeIII and 135-bp CCA-HaeIII fragments were composed of minisatellite-like and microsatellite-like repeats, respectively. ACH-Sau3AI was a homolog of the chicken nuclear membrane repeat sequence, whose homologs are common in Phasianidae. CVI-MspI, CCA-BamHI, and CSQ-BamHI showed high homology and were specific to the Odontophoridae. CVI-MspI was localized to microchromosomes, whereas CVI-HaeIII, CCA-BamHI, and CSQ-BamHI were mapped to almost all chromosomes. CCA-HaeIII was localized to five pairs of macrochromosomes and most microchromosomes. ACH-Sau3AI was distributed in three pairs of macrochromosomes and all microchromosomes. Centromeric repetitive sequences may be homogenized in chromosome size-correlated and -uncorrelated manners in New World quails, although there may be a mechanism that causes homogenization of centromeric repetitive sequences primarily between microchromosomes, which is commonly observed in phasianid birds.

A comparative study of genome organization and inferences for the systematics of two large bushcricket genera of the tribe Barbitistini (Orthoptera: Tettigoniidae: Phaneropterinae)

BackgroundPoecilimon and Isophya are the largest genera of the tribe Barbitistini and among the most systematically complicated and evolutionarily intriguing groups of Palearctic tettigoniids. We examined the genomic organization of 79 taxa with a stable chromosome number using classical (C–banding, silver and fluorochrome staining) and molecular (fluorescence in situ hybridization with 18S rDNA and (TTAGG) n telomeric probes) cytogenetic techniques. These tools were employed to establish genetic organization and differences or similarities between genera or species within the same genus and determine if cytogenetic markers can be used for identifying some taxonomic groups of species.ResultsDifferences between the karyotypes of the studied genera include some general changes in the morphology of the X chromosome in Isophya (in contrast to Poecilimon). The number of major rDNA clusters per haploid genome divided Poecilimon into two main almost equal groups (with either one or two clusters), while two rDNA clusters predominated in Isophya. In both genera, rDNA loci were preferentially located in the paracentromeric region of the autosomes and rarely in the sex chromosomes. Our results demonstrate a coincidence between the location of rDNA loci and active NORs and GC-rich heterochromatin regions. The C/DAPI/CMA3 bands observed in most Poecilimon chromosomes suggest the presence of more families of repetitive DNA sequences as compared to the heterochromatin patterns in Isophya.ConclusionsThe results show both differences and similarities in genome organization among species of the same genus and between genera. Previous views on the systematics and phylogenetic grouping of certain lineages are discussed in light of the present cytogenetic results. In some cases, variation of chromosome markers was observed to correspond with variation in other evolutionary traits, which is related to the processes of ongoing speciation and hybridization in zones of secondary contact. It was concluded that the physical mapping of rDNA sequences and heterochromatin may be used as an additional marker for understanding interspecific relationships in these groups and their routes of speciation.

Divergence of Repetitive DNA Sequences in the Heterochromatin of Medaka Fishes: Molecular Cytogenetic Characterization of Constitutive Heterochromatin in Two Medaka Species: Oryzias hubbsi and O. celebensis (Adrianichthyidae, Beloniformes)

The large biarmed chromosomes of Oryzias celebensis [2n = 36, fundamental arm number (FN) = 48] are considered to have resulted from fusions of acrocentric chromosomes in the ancestral karyotype of Oryzias, which is retained in O. hubbsi (2n = 48, FN = 48). To understand the molecular evolution of heterochromatin associated with karyotype reorganization in medaka fishes, we cloned 3 and 6 novel families of heterochromatin-related repetitive DNA sequences from O. hubbsi and O. celebensis, respectively, and characterized them using molecular cytogenetics. Two AT-rich repetitive sequences isolated from the genomic DNA of O. hubbsi, a 164-bp satellite DNA (OHU-RsaI-Scen) and a 177-bp telomere-specific repeat (OHU-RsaI-Stelo), were shown to be major components of the constitutive heterochromatin of centromeres and telomeres, respectively. A GC-rich 326-bp sequence, named OHU-AluI-M1, was colocalized with the 18S-28S ribosomal RNA gene cluster to a single autosomal pair of chromosomes and the W chromosome. In O. celebensis, 2 major satellite DNA sequences (the AT-rich 157-bp OCE-AluI-Scen sequence and the 186-bp OCE-HinfI-Scen sequence) were identified in the centromeric regions of almost all chromosomes. The 197-bp OCE-HinfI-S6 sequence was located in the centromeric and distal and/or interstitial heterochromatin of almost all chromosomes, and the 191-bp OCE-HinfI-S8 sequence was located in 6 pairs of chromosomes. Constitutive heterochromatin on the short arm of large submetacentric chromosome 5 was composed of at least 3 different repetitive sequences: the 171-bp OCE-AluI-S18 sequence, the 197-bp OCE-HinfI-S6 sequence and the 172-bp OCE-HinfI-S11 sequence. All families of repeated sequences showed no nucleotide sequence similarity with each other and high species-specificity among 7 different species. These results suggest that the heterochromatin of O. hubbsi and O. celebensis consists of various types of repetitive sequence and that the sequences evolved independently and were then amplified site-specifically in each lineage after karyotype reorganization occurred in the ancestral karyotype.

Genomic Subtraction Recovers Rye-Specific DNA Elements Enriched in the Rye Genome

Repetitive DNA sequence families have been identified in methylated relic DNAs of rye. This study sought to isolate rye genome-specific repetitive elements regardless of the level of methylation, using a genomic subtraction method. The total genomic DNAs of rye-chromosome-addition-wheat lines were cleaved to short fragments with a methylation-insensitive 4-bp cutter, MboI, and then common DNA sequences between rye and wheat were subtracted by annealing with excess wheat genomic DNA. Four classes of rye-specific repetitive elements were successfully isolated from both the methylated and non-methylated regions of the genome. Annealing of the DNA mixture at a ratio of the enzyme-restricted fragments:the sonicated fragments (1:3-1:5) was key to this success. Two classes of repetitive elements identified here belong to representative repetitive families: the tandem 350-family and the dispersed R173 family. Southern blot hybridization patterns of the two repetitive elements showed distinct fragments in methylation-insensitive EcoO109I digests, but continuous smear signals in the methylation-sensitive PstI and SalI digests, indicating that both of the known families are contained in the methylated regions. The subtelomeric tandem 350-family is organized by multimers of a 380-bp-core unit defined by the restriction enzyme EcoO109I. The other two repetitive element classes had new DNA sequences (444, 89 bp) and different core-unit sizes, as defined by methylation-sensitive enzymes. The EcoO109I recognition sites consisting of PyCCNGGPu-multi sequences existed with high frequency in the four types of rye repetitive families and might be a useful tool for studying the genomic organization and differentiation of this species.

Molecular cloning and characterization of the repetitive DNA sequences that comprise the constitutive heterochromatin of the A and B chromosomes of the Korean field mouse (Apodemus peninsulae, Muridae, Rodentia)

Three novel families of repetitive DNA sequences were molecularly cloned from the Korean field mouse (Apodemus peninsulae) and characterized by chromosome in-situ hybridization and filter hybridization. They were all localized to the centromeric regions of all autosomes and categorized into major satellite DNA, type I minor, and type II minor repetitive sequences. The type II minor repetitive sequence also hybridized interspersedly in the non-centromeric regions. The major satellite DNA sequence, which consisted of 30 bp elements, was organized in tandem arrays and constituted the majority of centromeric heterochromatin. Three families of repetitive sequences hybridized with B chromosomes in different patterns, suggesting that the B chromosomes of A. peninsulae were derived from A chromosomes and that the three repetitive sequences were amplified independently on each B chromosome. The minor repetitive sequences are present in the genomes of the other seven Apodemus species. In contrast, the major satellite DNA sequences that had a low sequence homology are present only in a few species. These results suggest that the major satellite DNA was amplified with base substitution in A. peninsulae after the divergence of the genus Apodemus from the common ancestor and that the B chromosomes of A. peninsulae might have a species-specific origin.

A novel family of repetitive DNA sequences amplified site-specifically on the W chromosomes in Neognathous birds

A novel family of repetitive DNA sequences was molecularly cloned from ApaI-digested genomic DNA of two Galliformes species, Japanese quail (Coturnix japonica) and guinea fowl (Numida meleagris), and characterized by chromosome in-situ hybridization and filter hybridization. Both the repeated sequence elements produced intensely painted signals on the W chromosomes, whereas they weakly hybridized to whole chromosomal regions as interspersed-type repetitive sequences. The repeated elements of the two species had high similarity of nucleotide sequences, and cross-hybridized to chromosomes of two other Galliformes species, chicken (Gallus gallus) and blue-breasted quail (Coturnix chinensis). The nucleotide sequences were conserved in three other orders of Neognathous birds, the Strigiformes, Gruiformes and Falconiformes, but not in Palaeognathous birds, the Struthioniformes and Tinamiformes, indicating that the repeated sequence elements were amplified on the W chromosomes in the lineage of Neognathous birds after the common ancestor diverged into the Palaeognathae and Neognathae. They are components of the W heterochromatin in Neognathous birds, and a good molecular cytogenetic marker for estimating the phylogenetic relationships and for clarifying the origin of the sex chromosome heterochromatin and the process of sex chromosome differentiation in birds.

New families of site-specific repetitive DNA sequences that comprise constitutive heterochromatin of the Syrian hamster (Mesocricetus auratus, Cricetinae, Rodentia)

We molecularly cloned new families of site-specific repetitive DNA sequences from BglII- and EcoRI-digested genomic DNA of the Syrian hamster (Mesocricetus auratus, Cricetrinae, Rodentia) and characterized them by chromosome in situ hybridization and filter hybridization. They were classified into six different types of repetitive DNA sequence families according to chromosomal distribution and genome organization. The hybridization patterns of the sequences were consistent with the distribution of C-positive bands and/or Hoechst-stained heterochromatin. The centromeric major satellite DNA and sex chromosome-specific and telomeric region-specific repetitive sequences were conserved in the same genus (Mesocricetus) but divergent in different genera. The chromosome-2-specific sequence was conserved in two genera, Mesocricetus and Cricetulus, and a low copy number of repetitive sequences on the heterochromatic chromosome arms were conserved in the subfamily Cricetinae but not in the subfamily Calomyscinae. By contrast, the other type of repetitive sequences on the heterochromatic chromosome arms, which had sequence similarities to a LINE sequence of rodents, was conserved through the three subfamilies, Cricetinae, Calomyscinae and Murinae. The nucleotide divergence of the repetitive sequences of heterochromatin was well correlated with the phylogenetic relationships of the Cricetinae species, and each sequence has been independently amplified and diverged in the same genome.

Down-regulation of 7SL RNA Expression and Impairment of Vesicular Protein Transport Pathways by Leishmania Infection of Macrophages

The parasitic protozoan Leishmania specifically manipulates the expression of host macrophage genes during initial interactions, as revealed by mRNA differential display reverse transcription-PCR and cDNA microarray analyses. The genes that are down-regulated in mouse (J774G8) or human (U937) macrophages upon exposure to Leishmania include small RNA transcripts from the short interspersed element sequences. Among the short interspersed element RNAs that are down-regulated is 7SL RNA, which is the RNA component of the signal recognition particle. Because the microbicidal functions of macrophages profoundly count on vesicular protein transport processes, down-regulation of 7SL RNA may be significant in the establishment of infection by Leishmania in macrophage phagolysosomes. To evaluate whether down-regulation of 7SL RNA results in inhibition of signal recognition particle-mediated vesicular protein transport processes, we have tested and found that the targeting of proteins to the endoplasmic reticulum and plasma membrane and the secretion of proteins by macrophages are compromised in Leishmania-infected J774G8 and U937 cells. Knocking down 7SL RNA using small interfering RNA mimicked the effect of exposure of macrophages to Leishmania. The overexpression of 7SL RNA in J774G8 or U937 cells made these cells resistant to Leishmania infection, suggesting the possible biological significance of down-regulation of 7SL RNA synthesis in the establishment of infection by Leishmania. We conclude that Leishmania down-regulates 7SL RNA in macrophages to manipulate the targeting of many proteins that use the vesicular transport pathway and thus favors its successful establishment of infection in macrophages.

Molecular and cytogenetic characterization of site-specific repetitive DNA sequences in the Chinese soft-shelled turtle (Pelodiscus sinensis, Trionychidae)

A novel family of repetitive DNA sequences that are components of constitutive heterochromatin were cloned from BglI-digested genomic DNA of the Chinese soft-shelled turtle (Pelodiscus sinensis, Trionychidae), and characterized by filter hybridization and chromosome in-situ hybridization. The BglI-family of repetitive sequences were classified into four types by their genome organization and chromosomal distribution as follows: the repeated sequences located on (1) two pairs of microchromosomes, (2) four pairs of microchromosomes,(3) about half the number of microchromosomes and (4) the interstitial region of the short arm of chromosome 2. The presence of microchromosome-specific repetitive sequences has also been reported in the Struthioniformes and Galliformes, suggesting that turtle chromosomes retain some similarity to the chromosome structure as well as the karyotypes of avian species.

Different patterns in molecular evolution of the Triticeae.

A huge part of the genomes of most Triticeae species is formed by different families of repetitive DNA sequences. In this paper the phylogenetic distribution of two major classes of the repeats, retrotransposons and tandemly organized DNA sequences, are considered and compared with the evolution of gene-rich regions and generally accepted Triticeae phylogenetic relationships. In Hordeum, LTR-containing retrotransposons are dispersed along the chromosomes and are consistent with the existing picture of the phylogeny of Hordeum. Another retrotransposon class, LINEs, have evolved independently from LTR-retrotransposons. Different retrotransposon classes appear to have competed for genome space during the evolution of Hordeum. Another class of repeats, tandemly organized DNA sequences, tends to cluster at the functionally important regions of chromosomes, centromeres and telomeres. The distribution of a number of tandem DNA families in Triticeae is not congruent with generally accepted phylogenetic relationships. While natural selection is the dominant factor determining the structure of genic regions we suggest that the contribution of random events is important in the evolution of repetitive DNA sequences. The interplay of stochastic processes, molecular drive, and selection determines the structure of chromosomal regions, notably at centromeres and telomeres, stabilizing and differentiating species-specific karyotypes. Thus, the evolution of these regions may occur largely independently of the evolution of gene-rich regions.

An SSR-based genetic linkage map for perennial ryegrass ( Lolium perenne L.).

A simple sequence repeat (SSR)-based linkage map has been constructed for perennial ryegrass ( Lolium perenne L.) using a one-way pseudo-testcross reference population. A total of 309 unique perennial ryegrass SSR (LPSSR) primer pairs showing efficient amplification were evaluated for genetic polymorphism, with 31% detecting segregating alleles. Ninety-three loci have been assigned to positions on seven linkage groups. The majority of the mapped loci are derived from cloned sequences containing (CA)(n)-type dinucleotide SSR arrays. A small number (7%) of primer pairs amplified fragments that mapped to more than one locus. The SSR locus data has been integrated with selected data for RFLP, AFLP and other loci mapped in the same population to produce a composite map containing 258 loci. The SSR loci cover 54% of the genetic map and show significant clustering around putative centromeric regions. BLASTN and BLASTX analysis of the sequences flanking mapped SSRs indicated that a majority (84%) are derived from non-genic sequences, with a small proportion corresponding to either known repetitive DNA sequence families or predicted genes. The mapped LPSSR loci provide the basis for linkage group assignment across multiple mapping populations.

Identification of genes that are associated with DNA repeats in prokaryotes.

Using in silico analysis we studied a novel family of repetitive DNA sequences that is present among both domains of the prokaryotes (Archaea and Bacteria), but absent from eukaryotes or viruses. This family is characterized by direct repeats, varying in size from 21 to 37 bp, interspaced by similarly sized non-repetitive sequences. To appreciate their characteri-stic structure, we will refer to this family as the clustered regularly interspaced short palindromic repeats (CRISPR). In most species with two or more CRISPR loci, these loci were flanked on one side by a common leader sequence of 300-500 b. The direct repeats and the leader sequences were conserved within a species, but dissimilar between species. The presence of multiple chromosomal CRISPR loci suggests that CRISPRs are mobile elements. Four CRISPR-associated (cas) genes were identified in CRISPR-containing prokaryotes that were absent from CRISPR-negative prokaryotes. The cas genes were invariably located adjacent to a CRISPR locus, indicating that the cas genes and CRISPR loci have a functional relationship. The cas3 gene showed motifs characteristic for helicases of the superfamily 2, and the cas4 gene showed motifs of the RecB family of exonucleases, suggesting that these genes are involved in DNA metabolism or gene expression. The spatial coherence of CRISPR and cas genes may stimulate new research on the genesis and biological role of these repeats and genes.

Molecular and cytogenetic characterization of highly repeated DNA sequences in the vole Microtus cabrerae.

The genus Microtus presents several species with extremely large sex chromosomes that contain large blocks of constitutive heterochromatin. Several cytogenetic and molecular studies of the repetitive sequences in species of the genus Microtus have demonstrated that the heterochromatin is highly heterogeneous. We have cloned and characterized a family of repetitive DNA sequences from M. cabrerae, a species with large heterochromatic blocks on the giant sex chromosomes. These repetitive sequences are 65.84% A-T rich, organized in tandem, with a 161-bp unit and are located on the centromeric region of autosomes and the X chromosome. In addition, this repetitive DNA is located throughout the entire heterochromatic block of the X chromosome and on three interstitial bands in the heterochromatic block of the Y chromosome. Comparative analysis of this family of repetitive sequences from three Microtus species revealed that the development of these sequences has occurred by concerted evolution. Our results support the hypothesis that the heterochromatic blocks from the sex chromosomes of different species are evolving independently and they probably have the genetic capacity to amplify and retain different satellite DNAs. For a topic related to the location of these repetitive DNA sequences on the Y chromosome of M. cabrerae, we propose a model to explain the origin of a length polymorphism previously described for this chromosome.

A family of dispersed repetitive DNA sequences in tobacco contain clusters of W-box elements recognized by pathogen-induced WRKY DNA-binding proteins

We have previously identified in tobacco ( Nicotiana tabacum) a group of pathogen- and salicylic acid (SA)-induced DNA-binding proteins that contain the highly conserved WRKY DNA-binding domain. In order to identify their potential target genes, we have isolated tobacco genomic DNA sequences that contain the consensus TTGAC(C/T) binding sites (W boxes) recognized by WRKY DNA-binding proteins. Surprisingly, sequence analysis of the 16 clones with the strongest binding affinities for the WRKY proteins indicated that they all belong to a family of dispersed repetitive DNA sequences with an approximate copy number of 10,000 per haploid tobacco genome. These repetitive DNA sequences contain a number of direct and inverted repeats, a feature commonly attributed to mobile genetic elements. BLAST search revealed that similar repetitive sequences are present in the promoters of the tobacco par/ str246C gene and the gene encoding a feedback-insensitive anthranilate synthase α-2 chain. Interestingly, the par/ str246C gene, which contains a W-box element in the repetitive sequence of its promoter, was induced dramatically in resistant tobacco plants after infection with tobacco mosaic virus. These results support that dispersed repetitive DNA sequences may serve as reservoirs for new functional cis-acting DNA elements that can be recruited through chromosomal rearrangement to participate in transcriptional regulation of nearby genes.

Characterization of a Highly Repetitive Sequence Conserved Among the North American Morone Species.

: A highly repetitive DNA sequence family from the genome of the North American Morone has been cloned and characterized. This family, first identified as a HindIII repetitive element, is composed of repeat units that range from 285 to 288 bp in length and comprise approximately 5.5% of the genome. The copy number of the repeat was estimated to be 1.85 x 10(5) per haploid genome set. Data from Southern blot analyses demonstrated that the HindIII repetitive element was tandemly organized. Sequence analysis of six cloned repeat monomers from each of the four North American Morone species, M. saxatilis, M. chrysops, M. americana, and M. mississippiensis, revealed a high degree of conservation of the monomeric unit. The intraspecific sequence variation ranged from 3.2% to 5.4%. A similar level of variation was detected between cloned monomers from the same individual, suggesting that most of the intraspecies variation may be due to variation among copies of the repeat. The interspecific sequence variation ranged from less than 4.6% between M. americana and M. mississippiensis to approximately 16% between the other Morone species pairs. Phylogenetic analysis of the repetitive element nucleotide sequences indicated that M. americana and M. mississippiensis were more closely related to each other than to any other pairs of Morone species. In addition, we reconstructed the Morone phylogeny using 22 previously described morphologic characters. Congruent relationships were obtained between both sets of data. The data suggest that the genus Morone is composed of two sets of sister taxa, M. saxatilis:M. chrysops and M. americana:M. mississippiensis.

Molecular Analysis of the Wheat Blast Population in Brazil with a Homolog of Retrotransposon MGR583.

We have identified a family of dispersed repetitive DNA sequences in the genome of a Magnaporthe grisea isolate from finger millet (Eleusine coracana). Southern blot analyses showed that this element is present in a moderate copy number (30-40 copies) in the genome of blast isolates from wheat (Triticum aestivum). DNA sequence data suggested that this element contains a region highly homologous to the reverse transcriptase domain of MGR583, a poly A-type retrotransposon. Using the reverse transcriptase domain of this element as a molecular probe, the genetic structure of the wheat blast population was examined. DNA fingerprinting analyses revealed that the wheat-infecting isolates constitute a separate, single lineage of their own, suggesting that they are derived from a single origin.

Polymorphisms and genomic organization of repetitive DNA from centromeric regions of Arabidopsis chromosomes.

A highly abundant repetitive DNA sequence family of Arabidopsis, AtCon, is composed of 178-bp tandemly repeated units and is located at the centromeres of all five chromosome pairs. Analysis of multiple copies of AtCon showed 95% conservation of nucleotides, with some alternative bases, and revealed two boxes, 30 and 24 bp long, that are 99% conserved. Sequences at the 3' end of these boxes showed similarity to yeast CDEI and human CENP-B DNA-protein binding motifs. When oligonucleotides from less conserved regions of AtCon were hybridized in situ and visualized by using primer extension, they were detected on specific chromosomes. When used for polymerase chain reaction with genomic DNA, single primers or primer pairs oriented in the same direction showed negligible amplification, indicating a head-to-tail repeat unit organization. Most primer pairs facing in opposite directions gave several strong bands corresponding to their positions within AtCon. However, consistent with the primer extension results, some primer pairs showed no amplification, indicating that there are chromosome-specific variants of AtCon. The results are significant because they elucidate the organization, mode of amplification, dispersion, and evolution of one of the major repeated sequence families of Arabidopsis. The evidence presented here suggests that AtCon, like human alpha satellites, plays a role in Arabidopsis centromere organization and function.

Polymorphisms and Genomic Organization of Repetitive DNA from Centromeric Regions of Arabidopsis Chromosomes

A highly abundant repetitive DNA sequence family of Arabidopsis, AtCon, is composed of 178-bp tandemly repeated units and is located at the centromeres of all five chromosome pairs. Analysis of multiple copies of AtCon showed 95% conservation of nucleotides, with some alternative bases, and revealed two boxes, 30 and 24 bp long, that are 99% conserved. Sequences at the 3' end of these boxes showed similarity to yeast CDEI and human CENP-B DNA-protein binding motifs. When oligonucleotides from less conserved regions of AtCon were hybridized in situ and visualized by using primer extension, they were detected on specific chromosomes. When used for polymerase chain reaction with genomic DNA, single primers or primer pairs oriented in the same direction showed negligible amplification, indicating a head-to-tail repeat unit organization. Most primer pairs facing in opposite directions gave several strong bands corresponding to their positions within AtCon. However, consistent with the primer extension results, some primer pairs showed no amplification, indicating that there are chromosome-specific variants of AtCon. The results are significant because they elucidate the organization, mode of amplification, dispersion, and evolution of one of the major repeated sequence families of Arabidopsis. The evidence presented here suggests that AtCon, like human alpha satellites, plays a role in Arabidopsis centromere organization and function.

Molecular phylogenetics of the hominoid Y chromosome.

The Human Y-chromosome plays a central role in sex determination, and is composed of DNA sequences homologous to the Y-chromosome, families of Y-specific repetitive DNA sequences, and single copy sequences. We investigated the chromosomal location of Y-specific DNA sequences, in the chimpanzee (Pan troglodytes), gorilla (Gorilla gorilla), and orangutan (Pongo pygmaeus) by the fluorescence in situ hybridization (FISH) technique. The Yq subtelomeric DNA sequences (DYS427) have been observed to be intact at the presumed loci. Also, the amelogenin gene (AMELY, Yp11.2) revealed sequence homology and positional conservation in the higher primates, except in gorilla where positional divergence was observed.