Repetitive DNA Clones Research Articles

Constructing an alternative wheat karyotype using barley genomic DNA.

The established karyotype was generated by genomic in situ hybridization (GISH) using total barley genomic DNA as labelled probe on mitotic metaphase bread wheat chromosomes. GISH produced specific banding signals on 16 of the 21 chromosome pairs. The following chromosomes showed distinctive banding patterns: 2A, 3A, 4A, 5A, 6A, 7A, 1D, 2D, 7D and all of the B chromosomes. The remaining chromosomes showed either faint bands or no hybridization signals at all. The in situ hybridization patterns corresponded to the GAA-satellite sequence, which is similar to the N-banding pattern in wheat. In situ hybridization by labelling total barley genomic DNA made it possible to identify most of the bread wheat chromosomes. The present paper describes a GISH-banding method for hexaploid wheat chromosomes. It is a valuable alternative method for fast chromosome selection without using FISH repetitive DNA clones.

Development of T. aestivum L.–H. californicum Alien Chromosome Lines and Assignment of Homoeologous Groups of Hordeum californicum Chromosomes

Hordeum californicum (2n = 2x = 14, HH) is resistant to several wheat diseases and tolerant to lower nitrogen. In this study, a molecular karyotype of H. californicum chromosomes in the Triticum aestivum L. cv. Chinese Spring (CS)–H. californicum amphidiploid (2n = 6x = 56, AABBDDHH) was established. By genomic in situ hybridization (GISH) and multicolor fluorescent in situ hybridization (FISH) using repetitive DNA clones (pTa71, pTa794 and pSc119.2) as probes, the H. californicum chromosomes could be differentiated from each other and from the wheat chromosomes unequivocally. Based on molecular karyotype and marker analyses, 12 wheat–alien chromosome lines, including four disomic addition lines (DAH1, DAH3, DAH5 and DAH6), five telosomic addition lines (MtH7L, MtH1S, MtH1L, DtH6S and DtH6L), one multiple addition line involving H. californicum chromosome H2, one disomic substitution line (DSH4) and one translocation line (TH7S/1BL), were identified from the progenies derived from the crosses of CS–H. californicum amphidiploid with common wheat varieties. A total of 482 EST (expressed sequence tag) or SSR (simple sequence repeat) markers specific for individual H. californicum chromosomes were identified, and 47, 50, 45, 49, 21, 51 and 40 markers were assigned to chromosomes H1, H2, H3, H4, H5, H6 and H7, respectively. According to the chromosome allocation of these markers, chromosomes H2, H3, H4, H5, and H7 of H. californicum have relationship with wheat homoeologous groups 5, 2, 6, 3, and 1, and hence could be designated as 5Hc, 2Hc, 6Hc, 3Hc and 1Hc, respectively. The chromosomes H1 and H6 were designated as 7Hc and 4Hc, respectively, by referring to SSR markers located on rye chromosomes.

Characterization of a leaf rust-resistant wheat–Thinopyrum ponticum partial amphiploid BE-1, using sequential multicolor GISH and FISH

In situ hybridization (multicolor GISH and FISH) was used to characterize the genomic composition of the wheat-Thinopyrum ponticum partial amphiploid BE-1. The amphiploid is a high-protein line having resistance to leaf rust (Puccinia recondita f. sp. tritici) and powdery mildew (Blumeria graminis f. sp. tritici) and has in total 56 chromosomes per cell. Multicolor GISH using J, A and D genomic probes showed 16 chromosomes originating from Thinopyrum ponticum and 14 A genome, 14 B genome and 12 D genome chromosomes. Six of the Th. ponticum chromosomes carried segments different from the J genome in their centromeric regions. It was demonstrated that these alien chromosome segments did not originate from the A, B or D genomes of wheat, so the translocation chromosomes were considered to be J(s) type chromosomes carrying segments similar to the S genome near the centromeres. Rearrangements between the A and D genomes of wheat were detected. FISH using Afa family, pSc119.2 and pTa71 probes allowed the identification of all the wheat chromosomes present and the determination of the chromosomes involved in the translocations. The 4A and 7A chromosomes were identified as being involved in intergenomic translocations. The replaced wheat chromosome was identified as 7D. The localization of these repetitive DNA clones on the Th. ponticum chromosomes of the amphiploid was described in the present study. On the basis of their multicolor FISH patterns, the alien chromosomes could be arranged in eight pairs and could also be differentiated unequivocally from each other.

Molecular cytogenetic characterization ofAegilops biuncialisand its use for the identification of 5 derived wheat – Aegilops biuncialisdisomic addition lines

The aim of the experiments was to produce and identify different Triticum aestivum-Aegilops biuncialis disomic addition lines. To facilitate the exact identification of the Ae. biuncialis chromosomes in these Triticum aestivum-Ae. biuncialis disomic additions, it was necessary to analyze the fluorescence in situ hybridization (FISH) pattern of Ae. biuncialis (2n = 4x = 28, U(b)U(b)M(b)M(b)), comparing it with the diploid progenitors (Aegilops umbellulata, 2n = 2x = 14, UU and Aegilops comosa, 2n = 2x = 14, MM). To identify the Ae. biuncialis chromosomes, FISH was carried out using 2 DNA clones (pSc119.2 and pAs1) on Ae. biuncialis and its 2 diploid progenitor species. Differences in the hybridization patterns of all chromosomes were observed among the 4 Ae. umbellulata accessions, the 4 Ae. comosa accessions, and the 3 Ae. biuncialis accessions analyzed. The hybridization pattern of the M genome was more variable than that of the U genome. Five different wheat-Ae. biuncialis addition lines were produced from the wheat-Ae. biuncialis amphiploids produced earlier in Martonvásár. The 2M, 3M, 7M, 3U, and 5U chromosome pairs were identified with FISH using 3 repetitive DNA clones (pSc119.2, pAs1, and pTa71) in the disomic chromosome additions produced. Genomic in situ hybridization (GISH) was used to differentiate the Ae. biuncialis chromosomes from wheat, but no chromosome rearrangements between wheat and Ae. biuncialis were detected in the addition lines.

A comparison of DNA- and RNA-based clone libraries from the same marine bacterioplankton community

Clones from the same marine bacterioplankton community were sequenced, 100 clones based on DNA (16S rRNA genes) and 100 clones based on RNA (16S rRNA). This bacterioplankton community was dominated by α- Proteobacteria in terms of repetitive DNA clones (52%), but γ- Proteobacteria dominated in terms of repetitive RNA clones (44%). The combined analysis led to a characterization of phylotypes otherwise uncharacterized if only the DNA or RNA libraries would have been analyzed alone. Of the DNA clones, 25.5% were found only in this library and no close relatives were detected in the RNA library. For clones from the RNA library, 21.5% of RNA clones did not indicate close relatives in the DNA library. Based on the comparisons between DNA and RNA libraries, our data indicate that the characterization of the bacterial community based on RNA has the potential to characterize distinct phylotypes from the marine environment, which remain undetected on the DNA level.

Fluorescence in situ hybridization polymorphism using two repetitive DNA clones in different cultivars of wheat

AbstractTwenty‐two wheat cultivars and a wheat line were analysed with two‐colour fluorescence in situ hybridization (FISH) using the pSc119.2 and pAs1 repetitive DNA clones to detect if polymorphism could be observed in the hybridization patterns of different wheat cultivars. The FISH hybridization pattern of ‘Chinese Spring’ was compared with wheat cultivars of different origins. Differences were observed in the hybridization patterns of chromosomes 4A, 5A, 1B, 2B, 3B, 5B, 6B, 7B, 1D, 2D, 3D and 4D. Although a low level of polymorphism exists in the FISH pattern of different wheat cultivars, it is possible to identify 17 pairs of chromosomes according to their hybridization patterns with these two probes. This study will help to predict the expected variation in the FISH pattern when analysing wheat genetic stocks of different origin. It is presumed that variation in hybridization patterns are caused by chromosome structural rearrangements and by differences in the amount and location of repetitive sequences in the cultivars analysed.

Isolation, characterization, and analysis of Leymus-specific DNA sequences.

Genomic Southern hybridization using labeled total genomic DNA of Leymus mollis as probe showed intense hybridization signals on all restriction enzyme digested DNA from five species of Leymus Hochst., and four species of Psathyrostachys Nevski. Experiments using the same L. mollis probe, but with unlabeled blocking DNA from Psathyrostachys, showed no hybridization at all. These two genera evidently had the same genomic content. Southern hybridization without blocking allowed identification of DNA fragments abundant in Leymus and Psathyrostachys. Fragments potentially specific to Leymus were cloned. Five repetitive DNA clones from L. mollis and L. arenarius were characterized: pLmIs1, pLmIs44, pLmIs51, pLmIs53, and pLaIs56. These clones hybridized to both Leymus and Psathyrostachys on Southern blots - no clone hybridized to only one of these genera. Both Southern blot and fluorescence in situ hybridization (FISH) experiments showed that all the clones contained dispersed repetitive sequences. They painted all and whole chromosomes uniformly except at centromeres, telomeres, and nucleolar organiser regions. Three of these clones, i.e., pLmIs1, pLmIs44, and pLmIs53, were essentially specific to Leymus and Psathyrostachys - little or no hybridization was detected in other genera such as Triticum, Hordeum, Thinopyrum, or Elymus. Sequence analysis further revealed that the clones were part of retroelements. In particular, the clone pLmIs44 produced hybridization profiles suitable for analysis of genetic relatedness among species. The present study shows that Leymus and Psathyrostachys share the same basic genome, Ns, and therefore provides strong evidence for combining these two genera.

Molecular cytogenetic analysis of wheat-alien hybrids and derivatives

New wheat × barley, wheat ×Aegilops biuncialisand wheat × rye hybrids were produced with the aim of alien gene transfer from these species into wheat. Amphiploids were produced with the help of colchicine treatment from the last two combinations. The new wheat × barley hybrids were multiplied in tissue culture because of the high degree of sterility and then pollinated with wheat to obtain backcross progenies. Wheat-barley chromosome pairing was detected using genomicin situhybridization (GISH) in two combinations (Mv9 kr1 × Igri, Asakazekomugi × Manas).In vitroconditions caused an increase in chromosome arm association frequency in both combinations and in fertility in some regenerants. Five wheat-barley translocations were produced in a wheat background and characterized through the combination of cytogenetic and molecular genetic approaches (GISH, FISH and SSR markers). The following translocations were identified: 2DS.2DL-1HS, 3HS.3BL, 6BS.6BL-4HL, 4D-5HS and 7DL.7DS-5HS. Physical mapping of the SSR markers on chromosomes 1H and 5H was carried out using the intragenomic and interspecific translocation breakpoints and the centromere as physical landmarks. Disomic wheat-Aegilops biuncialisadditions were produced after backcrossing the wheat-Ae. biuncialisamphiploids. Fluorescencein situhybridization (FISH) was carried out using two repetitive DNA clones (pSc119.2 and pAs1) onAe. biuncialisand its two diploid progenitor species to detect chromosome polymorphism. The 7M and 3M disomic chromosome additions were selected and five more lines still need to be characterized. The octoploid triticale (Mv9 kr1 × Lovászpatonai) produced in Martonvásár was crossed with a 1RS.1BL wheat cultivar Matador. GISH analysis detected pairing between the 1RS arm of the translocation chromosome and that of Lovászpatonai rye in 32 % of the pollen mother cells, making it possible to select recombinants from this combination. The new recombinants between the 1RS of Petkus and the 1RS of Lovászpatonai rye cultivars are being analysed with the help of microsatellite markers.

Interstitial colocalization of two cervid satellite DNAs involved in the genesis of the Indian muntjac karyotype.

A number of repetitive DNA clones were generated from PCR amplifications of Indian muntjac genomic DNA using primer sequences derived from a white tailed deer satellite II DNA sequence. One clone (Mmv-0.7) was characterized and shown to be a cervid satellite II DNA clone. Multiple colored FISH studies with cervid satellite I (C5) and this satellite II clone (Mmv-0.7) to Chinese muntjac metaphase chromosomes localized both satellite DNAs at the pericentromeric regions of all chromosomes except for chromosome 3 and the Y chromosome, whereas chromosome 3 exhibited pericentromeric satellite II DNA only. Where distinguishable, the pericentromeric satellite II signals appeared terminally oriented with respect to satellite I. Six pairs of Chinese muntjac autosomes had interstitial satellite I sites with four of these autosomal pairs (chromosomes 1, 2 and two other smaller autosomal pairs) also exhibiting interstitial satellite II signals. An interstitial site on the X chromosome was found to have satellite II signals. For the Indian muntjac chromosomes, FISH studies revealed a pericentromeric hybridization for satellites I and II as well as 27 distinct interstitial hybridization sites, each having at least one of the satellite DNAs. These data were used to more precisely define the chromosome fusion-associated breakpoints that presumably led to the formation of the present-day Indian muntjac karyotype. It further hints at the possibility that the Indian muntjac karyotype may have evolved directly from a 2n = 70 ancestral karyotype rather than from an intermediate 2n = 46 Chinese muntjac-like karyotype.

Cytological and molecular characterization of repetitive DNA sequences of Solanum brevidens and Solanum tuberosum.

The chromosomal distribution, copy numbers, and nucleotide sequences were determined for four repetitive DNA clones, pSB1 and pSB7 of Solanum brevidens and pST3 and pST10 of Solanum tuberosum. Using fluorescence in situ hybridization (FISH), pSB1 and pSB7 were localized near the telomeres and in some centromeric and interstitial sites of S. brevidens chromosomes, but not in S. tuberosum chromosomes, after high stringency washes. The clone pST3 showed signals in the telomeric areas of a few chromosomes in S. tuberosum, but signals were not detected in S. brevidens. All three repeated sequences (pSB1, pSB7, and pST3) were detected in chromosomal areas that are typically known to contain tandemly repeated sequences. The S. tuberosum clone pST10 did not show signals in either species even at low stringency conditions. The estimated copy numbers of the four clones were 1500, 6750, 300, and 400 for pSB1, pSB7, pST3, and pST10, respectively, in the corresponding haploid genomes (S. brevidens and S. tuberosum). The inserts of the four clones pSB1, pSB7, pST3, and pST10 were 322, 167, 845 and 121 bp, respectively. After sequencing, no significant sequence homologies were found among the four clones. A homology search in sequence data bases showed that pSB7 has variable homology (78-100%) with another repetitive sequence of S. brevidens Sb4/2 depending on its subrepeat. It also showed some homology with one repeat of tomato (pLEG15) and one repeat of Solanum circaeifolium (pSC15).

Cytogenetics for the model system Arabidopsis thaliana.

A detailed karyotype of Arabidopsis thaliana is presented using meiotic pachytene cells in combination with fluorescence in situ hybridization. The lengths of the five pachytene bivalents varied between 50 and 80 microns, which is 20-25 times longer than mitotic metaphase chromosomes. The analysis confirms that the two longest chromosomes (1 and 5) are metacentric and the two shortest chromosomes (2 and 4) are acrocentric and carry NORs subterminally in their short arms, while chromosome 3 is submetacentric and medium sized. Detailed mapping of the centromere position further revealed that the length variation between the pachytene bivalents comes from the short arms. Individual chromosomes were unambiguously identified by their combinations of relative lengths, arm-ratios, presence of NOR knobs and FISH signals with a 5S rDNA probe and chromosome specific DNA probes. Polymorphisms were found among six ecotypes with respect to the number and map positions of 5S rDNA loci. All ecotypes contain 5S rDNA in the short arms of chromosomes 4 and 5. Three different patterns were observed regarding the presence and position of a 5S rDNA locus on chromosome 3. Repetitive DNA clones enabled us to subdivide the pericentromeric heterochromatin into a central domain, characterized by pAL1 and 106B repeats, which accommodate the functional centromere and two flanking domains, characterized by the 17 A20 repeat sequences. The upper flanking domains of chromosomes 4 and 5, and in some ecotypes also chromosome 3, contain a 5S rDNA locus. The detection of unique cosmids and YAC sequences demonstrates that detailed physical mapping of Arabidopsis chromosomes by cytogenetic techniques is feasible. Together with the presented karyotype this makes Arabidopsis a model system for detailed cytogenetic mapping.

Fluorescence in situ hybridization of potato somatohaploids and their somatic hybrid donors using two Solanum brevidens specific sequences

Two Solanum brevidens specific repetitive DNA clones (pSB1 and pSB7) were used simultaneously as probes in fluorescence in situ hybridization (FISH) for cytological studies of somatohaploids and their somatic hybrid donors. pSB1 was labelled with digoxigenin-11-dUTP and pSB7 was labelled with biotin-14-dATP and they were detected with reporter molecules conjugated to fluorescent dyes using digital imaging. The tandemly repeated sequences hybridized mostly near the telomeres of the chromosomes of S. brevidens. Using these two probes, it was possible to identify chromosomes containing repetitive DNA of S. brevidens both in the somatic hybrids between S. brevidens and S. tuberosum, and somatohaploids derived from the somatic hybrids. These cytological analyses showed that for the largest part genomes of the hexaploid somatic hybrids and their anther-derived triploid somatohaploids were composed of the genome of S. brevidens.

Evolutionary conservation and molecular characteristics of repetitive sequences of Drosophila koepferae

Thirteen middle repetitive DNA clones obtained from the genome of Drosophila koepferae have been tested for their evolutionary conservation in the other seven species of the buzzatii and martensis clusters (repleta group). All but two of these clones exhibit qualitatively similar patterns of hybridization in the eight species. The average interspecific hybridization signal is 85 per cent of that found intraspecifically, ranging from 73 to 93 per cent. Partial sequencing of six of these clones has shown sequences related to the retrotransposon Gypsy, first characterized in D. melanogaster, as well as to the Anopheles gambiae LINE elements T1Ag and Q. A fragment of a hitherto unknown, short inverted repeat transposable element has also been found. The evolutionary conservation of repetitive D. koepferae sequences seems to be related to the high proportion of simple DNA and inactive mobile elements in the genome of this species.

Isolation and characterization of a species-specific multicopy DNA sequence fromEntamoeba histolytica

A genomic library of Entamoeba histolytica (pathogenic strain HM-1:IMSS) was screened to detect repetitive DNA clones other than those from the highly abundant ribosomal DNA (rDNA). One such clone (HMc) had a 2.3 kb insert which hybridized with the main genome and not the rDNA circle. Southern hybridization of E. histolytica genomic DNA, digested with EcoR I and probed with HMc, showed multiple bands. The banding pattern was identical in all axenic pathogenic strains tested. Differences, however, existed when the banding pattern of a pathogenic strain was compared with that of a non-pathogenic strain. HMc was present in about 25-30 copies per genome in strain HM-1:IMSS. Nucleotide sequence analysis of HMc revealed a partial open reading frame which hybridized with a 1.35 kb poly A+ transcript in Northern blots. The deduced amino acid sequence did not, however, show significant homology with known proteins. The HMc sequence was found only in E. histolytica as it hybridized with 5 different axenic strains of E. histolytica but did not recognize other closely related species of Entamoeba. It has thus the potential to be used as a species-specific DNA probe.

Repeated DNA Sequence 350bp Family Cloned from Agropyron intermedium for Identification of the Agropyron Chromosomes Added to Common Wheat Lines.

Repeated DNA sequences cloned from Agropyron intermedium (2n =42, E1E1E2E2XX) enabled to identify the Ag. intermedium chromosomes in the common wheat genome by Southern and in situ hybridization. Four repetitive DNA clones that did not hybridize to the total genomic DNA of Triticum aestivum cv. Chinese Spring (CS) were screened from the Mbol-digested genomic DNA Iibrary of Ag. intermedium.

Repetitive DNA sequences from polyploid Elymus trachycaulus and the diploid progenitor species: detection and genomic affinity of Elymus chromatin added to wheat

A set of four repetitive DNA clones, pEt1, pEt2, pCb1, and pCb3, were isolated from SH-genome polyploid Elymus trachycaulus and H-genome diploid Critesion bogdanii. The clone Et1 represents a tandemly arranged telomeric sequence. Et2 represents tandem repeats interspersed along the entire length of individual chromosomes. The Cb1 sequence was more evenly dispersed. The Et1 clone shared homology with a 350 base pair family of rye sequences. The Cb3 sequence was evenly distributed in S- and H-genome species. All the repetitive DNA sequences were excellent markers for the specific detection and genomic affinity of Elymus chromatin added to wheat. All clones showed intragenomic variation in copy number and chromosomal location. Based on the analysis of this variation, we conclude that E. trachycaulus most probably originated from putative diploid H- and S-genome species resembling Critesion californicum and Pseudoroegneria spicata, respectively.Key words: wheatgrass, wheat–Elymus hybrid, addition lines, polyploidy, restriction fragment length polymorphism.

New evidence for tandem chromosome fusions in the karyotypic evolution of Asian muntjacs

A clone of highly repetitive DNA, designated C5, was isolated from DNA of female Chinese muntjac cells. The nucleotide sequence of this clone is 80%-85% homologous to that of the satellite IA clone and other highly repetitive DNA clones previously obtained from the Indian muntjac. Using C5 as a probe for in situ hybridizations to chromosome preparations of cells of both the Chinese and Indian muntjacs, we were able to show that these repeated sequences occur in centromeric heterochromatin of the chromosomes of both Chinese and Indian muntjac species. More significantly, non-random clusters of hybridization signals were detected on the arms of chromosomes of the Indian muntjac. These latter hybridization sites are postulated to be regions of interstitial heterochromatin and could be the remnants of centromeric heterochromatin from ancestral Chinese muntjac chromosomes. Our observations provide new supportive evidence for the tandem chromosome fusion theory that has been proposed for the evolution of the Indian muntjac karyotype.

Cloning and characterization of a species-specific repetitive DNA sequence from Loa loa

A genomic DNA library of Loa loa was constructed in λgt11 using EcoRI-digested DNA from microfilariae isolated from two West African patients. Screening with labeled L. loa DNA yielded several potential repetitive DNA clones. An MboI fragment of one of these, LL3M9, was identified and characterized. Sequence analysis of LL3M9 revealed an 839-bp fragment with an unusual 356-bp region containing 37 copies of the hexamer CTTAGG, many of which are arranged in repeated motifs of 12, 27 and 63 bp. This region shares many of the characteristics of eukaryotic satellite DNA. A synthetic oligonucleotide corresponding to the 27-bp repeated motif, LL3M9REP, was found to be both sensitive and species-specific by dot hybridization. Species specificity of LL3M9REP was confirmed by amplification of the repetitive region using genomic DNA as a template in the polymerase chain reaction.

Possible repetitive DNA markers for Eusorghum and Parasorghum and their potential use in examining phylogenetic hypotheses on the origin of Sorghum species

Genomic structures of two major species in section Eusorghum (Sorghum), Sorghum bicolor and Sorghum halepense, and their phylogenetic relationships with a species in section Parasorghum, Sorghum versicolor, were studied by using cloned repetitive DNA sequences from the three species. Of the five repetitive DNA clones isolated from S. bicolor and S. halepense, four produced qualitatively similar hybridization patterns with detectable variations in copy numbers of some of the restriction fragments on the Southern blots of the two genomic DNAs. One clone was shown to be diagnostic for S. halepense. Molecular analysis at the DNA level indicates that S. bicolor and S. halepense have similar but not identical genomes, consonant with differences in karyotypes, meiotic chromosome behaviors, morphology, and physiology of the species. In addition to five repetitive clones isolated from S. bicolor and S. halepense, eight more sequences were cloned from S. versicolor. Nine clones were found to be specific for either S. bicolor and S. halepense or S. versicolor. The remaining four had a moderate to strong homology with sequences present in all Sorghum species studied. We speculate that the genome in the common ancestor of Sorghum has differentiated to give rise to genomes of at least three major chromosome sizes; large, medium, and small, as seen at present. Amplifications, eliminations, rearrangements, and new syntheses of repetitive sequences may have been involved in genome differentiation of these species. The results also suggest that the S. versicolor genome has strongly diverged from the genomes of the two species in section Eusorghum.

Characterization of human heterochromatin by in situ hybridization with satellite DNA clones.

Biotinylated DNA from two satellite-related, repetitive DNA clones, pHuR 98 and pHuR 195 (specific for chromosomes 9 and 16, respectively), and from a Y-specific clone, pY-3.4A, were hybridized to human metaphase chromosomes using fluoresceinated avidin to detect binding. The chromosomes were simultaneously counterstained with distamycin-DAPI to identify the AT-rich heterochromatin of chromosomes 1, 9, 15, 16, and the Y chromosome. With this method, clear results were obtained under both normal and low stringency conditions, allowing hybridization between molecules sharing 80-85% and 60-65% identity, respectively. Thus, additional sites related to the probes could be identified. A close relationship was shown between the heterochromatin of chromosomes 1 and 16, both hybridizing with clone pHuR 195 under low stringency. Hybridization with clone pHuR 98 was highly specific for chromosome 9, even under low stringency. A relationship between chromosomes 9, 15, and the Y chromosome, however, was shown by hybridization with clone pY-3.4A. The chromosomal distribution of the three repetitive DNA clones used in this study, and data from the literature, are in accordance with the distribution of the heterochromatin types characterized by staining with different fluorescent dyes and dye combinations. Furthermore, our sequence data for clones pHuR 98 and pHuR 195 may explain the fluorescent properties on which the cytogenetic classification of the heterochromatin is based.