Nucleotide sequence of BamHI family satellite DNA and its unit length polymorphism in bluegill sunfish Lepomis macrochirus

We have isolated and sequenced a member of tandem repetitive DNA containing BamHI site (BamHI family satellite DNA) from bluegill sunfish Lepomis macrochirus. PCR amplification with specific primers was performed to define the size of unit length repeat of the BamHI family satellite DNA, revealing that there were two distinct size of DNA fragments (0.9 kb and 1.3 kb) in the PCR products. The longer fragment (1.3 kb) consisted of internal sub-duplication of shorter fragment (0.9 kb). We have compared the size of PCR products among four fish populations, and found that both fragments co-existed in one population whereas the longer fragment was dominant in other three populations. The results may reflect ongoing homogenization of satellite DNA type over a short evolutionary time scale.     

Satellite DNA and chromosomes in Neotropical fishes: methods,applications and perspectives

Constitutive heterochromatin represents a substantial portion of the eukaryote genome, and it is mainly composed of tandemly repeated DNA sequences, such as satellite DNAs, which are also enriched by other dispersed repeated elements, including transposons. Studies on the organization, structure, composition and in situ localization of satellite DNAs have led to consistent advances in the understanding of the genome evolution of species, with a particular focus on heterochromatic domains, the diversification of heteromorphic sex chromosomes and the origin and maintenance of B chromosomes. Satellite DNAs can be chromosome specific or species specific, or they can characterize different species from a genus, family or even representatives of a given order. In some cases, the presence of these repeated elements in members of a single clade has enabled inferences of a phylogenetic nature. Genomic DNA restriction, using specific enzymes, is the most frequently used method for isolating satellite DNAs. Recent methods such as C₀t–1 DNA and chromosome microdissection, however, have proven to be efficient alternatives for the study of this class of DNA. Neotropical ichthyofauna is extremely rich and diverse enabling multiple approaches with regard to the differentiation and evolution of the genome. Genome components of some species and genera have been isolated, mapped and correlated with possible functions and structures of the chromosomes. The 5SHindIII‐DNA satellite DNA, which is specific to Hoplias malabaricus of the Erythrinidae family, has an exclusively centromeric location. The As51 satellite DNA, which is closely correlated with the genome diversification of some species from the genus Astyanax, has also been used to infer relationships between species. In the Prochilodontidae family, two repetitive DNA sequences were mapped on the chromosomes, and the SATH 1 satellite DNA is associated with the origin of heterochromatic B chromosomes in Prochilodus lineatus. Among species of the genus Characidium and the Parodontidae family, amplifications of satellite DNAs have demonstrated that these sequences are related to the differentiation of heteromorphic sex chromosomes. The possible elimination of satellite DNA units could explain the genome compaction that occurs among some species of Neotropical Tetraodontiformes. These topics are discussed in the present review, showing the importance of satellite DNA analysis in the differentiation and karyotype evolution of Actinopterygii.     

Structure Analysis of Two <Emphasis Type="Italic">Toxoplasma gondii</Emphasis> and <Emphasis Type="BoldItalic">Neospora caninum</Emphasis> Satellite DNA Families and Evolution of Their Common Monomeric Sequence

A family of repetitive DNA elements of approximately 350 bp—Sat350—that are members of Toxoplasma gondii satellite DNA was further analyzed. Sequence analysis identified at least three distinct repeat types within this family, called types A, B, and C. B repeats were divided into the subtypes B1 and B2. A search for internal repetitions within this family permitted the identification of conserved regions and the design of PCR primers that amplify almost all these repetitive elements. These primers amplified the expected 350-bp repeats and a novel 680-bp repetitive element (Sat680) related to this family. Two additional tandemly repeated high-order structures corresponding to this satellite DNA family were found by searching the Toxoplasma genome database with these sequences. These studies were confirmed by sequence analysis and identified: (1) an arrangement of AB1CB2 350-bp repeats and (2) an arrangement of two 350-bp-like repeats, resulting in a 680-bp monomer. Sequence comparison and phylogenetic analysis indicated that both high-order structures may have originated from the same ancestral 350-bp repeat. PCR amplification, sequence analysis and Southern blot showed that similar high-order structures were also found in the Toxoplasma-sister taxon Neospora caninum. The Toxoplasma genome database ( ) permitted the assembly of a contig harboring Sat350 elements at one end and a long nonrepetitive DNA sequence flanking this satellite DNA. The region bordering the Sat350 repeats contained two differentially expressed sequence-related regions and interstitial telomeric sequences.     

<Emphasis Type="Italic">Pearl</Emphasis>, a Novel Family of Putative Transposable Elements in Bivalve Mollusks

Abstract While genome sequencing projects have discovered numerous types of transposable elements in diverse eukaryotes, there are many taxa of ecological and evolutionary significance that have received little attention, such as the molluscan class Bivalvia. Examination of a 0.7-MB genomic sequence database from the cupped oyster Crassostrea virginica revealed the presence of a common interspersed element, CvA. CvA possesses subterminal inverted repeats, a tandemly repeated core element, a tetranucleotide microsatellite region, and the ability to form stable secondary structures. Three other less abundant repetitive elements with a similar structure but little sequence similarity were also found in C. virginica. Ana-1, a repetitive element with similar features, was discovered in the blood ark Anadara trapezia by probing a genomic library with a dimeric repeat element contained in intron 2 of a minor globin gene in that species. All of these elements are flanked by the dinucleotide AA, a putative target-site duplication. They exhibit structural similarity to the sea urchin Tsp family and Drosophila SGM insertion sequences; in addition, they possess regions of sequence similarity to satellite DNA from several bivalve species. We suggest that the Crassostrea repetitive elements and Ana-1 are members of a new MITE-like family of nonautonomous transposable elements, named pearl. Pearl is the first putative nonautonomous DNA transposon to be identified in the phylum Mollusca.     

Selected nuclear LINE elements with mitochondrial-DNA-like inserts are more plentiful and mobile in tumor than in normal tissue of mouse and rat

The nuclear DNA of normal and tumor mouse and rat tissue was examined for mitochondrial-DNA-like inserts by means of the Southern blot technique. The two probes were ³²P-labeled cloned mitochondrial DNA. KpnI, which doesn't cut either mitochondrial DNA, was one of the restriction enzymes, while the enzymes that fragment mitochondrial DNA were for mouse and rat PstI and BamHI, respectively. When KpnI alone was used in the procedure a nuclear LINE family whose elements had mitochondrial-DNA-like insertions was selected. Such elements were much more abundant in tumor than in normal tissue. The results with PstI alone and BamHI alone and each combined with KpnI indicated that there were mobile LINE elements with mitochondrial-DNA-like inserts in the nuclear genome of tumor. The mouse tissues were normal liver and a transplantable lymphoid leukemic ascites cell line L1210 that had been carried for 40 years. The rat tissues were normal liver and a hepatoma freshly induced by diethylnitrosoamine in order to minimize the role of 40 years of transplantation. Our unitary hypothesis for carcinogenesis of 1971, which suggested these experiments, has been augmented to include mobile nuclear elements with inserts of mitochondrial-DNA-like sequences. Such elements have been related to diseases of genetic predisposition such as breast cancer and Huntington's disease. J. Cell. Biochem. 68:100–109, 1998. © 1998 Wiley-Liss, Inc.     

Two different satellite DNAs in Beta vulgaris L.: evolution,quantification and distribution in the genus

Summary Two highly repeated EcoRI (0.45 × 10⁶) and BamHI (0.17 × 10⁶) fragments per haploid genome were found in sugar beet genomic DNA. Both fragments were located by 6% acrylamide-gel electrophoresis, purified and cloned in pUC18. Four of the inserts corresponding to each family were chosen for further study. Both fragment families display the main characteristics of the satellite DNA of animals and plants. The EcoRI and BamHI fragment families are arranged in long tandem arrays. Fragments of the EcoRI family (pBVE) were analyzed. They vary both in sequence and in length (158–160 nt) in comparison with the consensus sequence of 159nt. Both families are A-T rich; pBVE is 59% rich while pBVB is 69% rich. The BVESAT family is present in all the members of the section Vulgares. It is conserved in the section Procumbentes with 80% homology and the same length, but is not detectable in the Corollinae. The sequence variation rate and the variation in length (330±5 nt) are of the same order in comparison with those of the BVESAT family. However, the BVBSAT family is present in species of the section Vulgares only. As regards other plant satellite DNAs, the BVESAT family shares homology with Allium cepa satellite DNA, with three of the yeast centromeric sequences, and with three Arabidopsis thaliana sequences. The BVBSAT family is unique to the Vulgares and does not share any homology with other plant or animal satellite DNAs sequences so far.     

Complex Evolution of Tandem-Repetitive DNA in the Chironomus thummi Species Group

The subspecies Chironomus thummi thummi and C. t. piger display dramatic differences in the copy number and chromosomal localization of a tandemly repeated DNA family (Cla elements). In order to analyze the evolutionary dynamics of this repeat family, we studied the organization of Cla elements in the related outgroup species C. luridus. We find three different patterns of Cla element organization in C. luridus, showing that Cla elements may be either strictly tandem-repetitive or be an integral part of two higher-order tandem repeats (i.e., Hinf[lur] elements, Sal[lur] elements). All three types of Cla-related repeats are localized in the centromeres of C. luridus chromosomes. This suggests that the dispersed chromosomal localization of Cla elements in C. t. thummi may be the result of an amplification and transposition during evolution of this subspecies. Received: 22 May 1996 / Accepted: 8 October 1996     

Interhomologue sequence variation of alpha satellite DNA from human chromosome 17: Evidence for concerted evolution along haplotypic lineages

Alpha satellite DNA is a family of tandemly repeated DNA found at the centromeres of all primate chromosomes. Different human chromosomes 17 in the population are characterized by distinct alpha satellite haplotypes, distinguished by the presence of variant repeat forms that have precise monomeric deletions. Pairwise comparisons of sequence diversity between variant repeat units from each haplotype show that they are closely related in sequence. Direct sequencing of PCR-amplified alpha satellite reveals heterogeneous positions between the repeat units on a chromosome as two bands at the same position on a sequencing ladder. No variation was detected in the sequence and location of these heterogeneous positions between chromosomes 17 from the same haplotype, but distinct patterns of variation were detected between chromosomes from different haplotypes. Subsequent sequence analysis of individual repeats from each haplotype confirmed the presence of extensive haplotype-specific sequence variation. Phylogenetic inference yielded a tree that suggests these chromosome 17 repeat units evolve principally along haplotypic lineages. These studies allow insight into the relative rates and/or timing of genetic turnover processes that lead to the homogenization of tandem DNA families. Correspondence to: H.F. Willard     

Sequence analysis of Vicia faba highly repeated DNA: the BamHI repeated sequence families

Cleavage of Vicia faba nuclear DNA with the restriction endonuclease BamHI yielded discrete size classes of 250, 850, 900, 990, 1 150, 1 500 and 1 750 bp of highly repetitive DNA. Each of these sequence families comprised about 3% of the total genomic DNA. Some sequence members from each sequence family were cloned in pBR322 and their primary structures determined. Computer analyses of nucleotide sequences suggested the existence of about 60 bp sequence periodicity within the repeating unit of the 990 bp sequence family, though the extent of homology among the surmised shorter subrepeat units was very low. With other BamHI sequence families, however, the data did not show any clear internal sequence periodicity. The repeat units of the 850 bp and 1 750 bp sequence families contained nucleotide sequences homologous to the 250 bp family sequence. No sequence relationship between or among other sequence families was observed. There was 13–25% sequence variation among 6 cloned members of the 250 bp family and probably also among those of other BamHI repeat families. DNA sequences homologous to these V. faba BamHI repeat families were detected in Pisum sativum DNA by Southern blot hybridization. Furthermore, very weak cross-hybridization was observed with plant DNAs from Phaseolus vulgaris, Triticum aestivum, Cucumis sativus and Trillium kamtschaticum.     

Evolution of Tandemly Repeated Sequences: What Happens at the End of an Array?

Tandemly repeated sequences are a major component of the eukaryotic genome. Although the general characteristics of tandem repeats have been well documented, the processes involved in their origin and maintenance remain unknown. In this study, a region on the paternal sex ratio (PSR) chromosome was analyzed to investigate the mechanisms of tandem repeat evolution. The region contains a junction between a tandem array of PSR2 repeats and a copy of the retrotransposon NATE, with other dispersed repeats (putative mobile elements) on the other side of the element. Little similarity was detected between the sequence of PSR2 and the region of NATE flanking the array, indicating that the PSR2 repeat did not originate from the underlying NATE sequence. However, a short region of sequence similarity (11/15 bp) and an inverted region of sequence identity (8 bp) are present on either side of the junction. These short sequences may have facilitated nonhomologous recombination between NATE and PSR2, resulting in the formation of the junction. Adjacent to the junction, the three most terminal repeats in the PSR2 array exhibited a higher sequence divergence relative to internal repeats, which is consistent with a theoretical prediction of the unequal exchange model for tandem repeat evolution. Other NATE insertion sites were characterized which show proximity to both tandem repeats and complex DNAs containing additional dispersed repeats. An ``accretion model' is proposed to account for this association by the accumulation of mobile elements at the ends of tandem arrays and into ``islands' within arrays. Mobile elements inserting into arrays will tend to migrate into islands and to array ends, due to the turnover in the number of intervening repeats. Received: 18 August 1997 / Accepted: 18 September 1998     

Conservation of a highly repeated DNA family of Aedes albopictus among mosquito genomes (Diptera: Culicidae)

Summary The genomic organization and chromosomal localization of a cloned 0.79-kb highly repeated DNA fragment, H-115, isolated from Aedes albopictus has been examined. The cloned fragment is a part of a larger unit of 1.86 kb that is tandemly repeated in the Ae. albopictus genome. The H-115 family of sequences are located at the intercalary position on chromosome 1 in Ae. albopictus. Similar patterns of in situ and Southern blot hybridization results are obtained in Ae. aegypti, Ae. seatoi, Ae. flavopictus, Ae. polynesiensis, Ae. Alcasidi, and Ae. katherinensis. The H-115 sequences are widely conserved in Culicidae and are found in Haemagogus equinus, Tripteroides bambusa, and Anopheles quadrimaculatus by hybridization under high stringency conditions. The H-115 sequences are also tandemly repeated in Hg. equinus with a monomer unit of 1.86 kb and in Tp. bambusa with a slightly diverged monomer unit of 1.90kb. In Anopheles quadrimaculatus, the H-115 sequences are dispersed throughout the genome. Partial sequence analysis shows that the H-115 insert is 62% AT and contains two perfect inverted repeats and numerous perfect direct repeats. The occurrence of inverted repeats with potential to form intrastrand palindromic structure suggests that the H-115 family of sequences may be involved in chromatin condensation.     

The organisation,nucleotide sequence,and chromosomal distribution of a satellite DNA from Allium cepa

We have investigated the organisation, nucleotide sequence, and chromosomal distribution of a tandemly repeated, satellite DNA from Allium cepa (Liliaceae). The satellite, which constitutes about 4% of the A. cepa genome, may be resolved from main-band DNA in antibiotic-CsCl density gradients, and has a repeat length of about 375 base pairs (bp). A cloned member of the repeat family hybridises exclusively to chromosome telomeres and has a non-random distribution in interphase nuclei. We present the nucleotide sequences of three repeats, which differ at a large number of positions. In addition to arrays made up of 375-bp repeats, homologous sequences are found in units with a greater repeat length. This divergence between repeats reflects the heterogeneity of the satellite determined using other criteria. Possible constraints on the interchromosomal exchange of repeated sequences are discussed.     

A molecular and cytogenetic survey of major repeated DNA sequences in tomato (Lycopersicon esculentum)

Summary The major families of repeated DNA sequences in the genome of tomato (Lycopersicon esculentum) were isolated from a sheared DNA library. One thousand clones, representing one million base pairs, or 0.15% of the genome, were surveyed for repeated DNA sequences by hybridization to total nuclear DNA. Four major repeat classes were identified and characterized with respect to copy number, chromosomal localization by in situ hybridization, and evolution in the family Solanaceae. The most highly repeated sequence, with approximately 77000 copies, consists of a 162 bp tandemly repeated satellite DNA. This repeat is clustered at or near the telomeres of most chromosomes and also at the centromeres and interstitial sites of a few chromosomes. Another family of tandemly repeated sequences consists of the genes coding for the 45 S ribosomal RNA. The 9.1 kb repeating unit in L. esculentum was estimated to be present in approximately 2300 copies. The single locus, previously mapped using restriction fragment length polymorphisms, was shown by in situ hybridization as a very intense signal at the end of chromosome 2. The third family of repeated sequences was interspersed throughout nearly all chromosomes with an average of 133 kb between elements. The total copy number in the genome is approximately 4200. The fourth class consists of another interspersed repeat showing clustering at or near the centromeres in several chromosomes. This repeat had a copy number of approximately 2100. Sequences homologous to the 45 S ribosomal DNA showed cross-hybridization to DNA from all solanaceous species examined including potato, Datura, Petunia, tobacco and pepper. In contrast, with the exception of one class of interspersed repeats which is present in potato, all other repetitive sequences appear to be limited to the crossing-range of tomato. These results, along with those from a companion paper (Zamir and Tanksley 1988), indicate that tomato possesses few highly repetitive DNA sequences and those that do exist are evolving at a rate higher than most other genomic sequences.     

Restriction site periodicities in highly repetitive DNA of primates.

Highly repeated DNA sequences from three Old World primate groups have been compared, using restriction endonucleases. Baboons, macaques and mangabeys share a 340⁴ base-pair, tandemly repeated DNA that is cut once by EndoR · BamHI. The several species of guenons, including the African green monkey, possess a related 170 base-pair, tandemly organized sequence distinguished by the feature of being cut once by EndoR · HindIII, EndoR · MboII or EndoR · HphI. The tandemly repeated DNA of the colobus monkey is based on a monomer length of 680 base-pairs, being cut once by EndoR · BamI or EndoR · EcoRI. Thus, all three highly repeated DNAs have a monomer length of 170n base-pairs, where n = 1, 2 or 4. The 340 and 680 base-pair repeated DNAs contain an internal 170 base-pair periodicity with respect especially to the EndoR · HindIII cleavage site, but with respect also to several other enzymes that characterize each repeated sequence. The 170 base-pair length is called the fundamental unit.The three repeated DNAs are more conserved in the region around the HindIII site and are more divergent elsewhere in the sequence. All seven 170 base-pair fundamental units were related to one another, judging from the overall similarities of the maps of restriction endonuclease cleavage sites. The highly repeated DNAs from baboons and guenons are related enough to cross-hybridize at relaxed criteria (60 °C in 0.12 m-Na⁺) but neither hybridizes to repeated colobus DNA under this condition.The results show that highly repeated sequences in primates form a common library descended from a single ancestral sequence, with 170 base-pairs making up the fundamental unit of library members. Occasionally, a member of the library is amplified, creating a newly amplified family. In Old World monkeys the most recent amplification just preceded active speciation.     

Organization of a dispersed repeated DNA element in the Zamia genome

Occurrence and genomic organization of dispersed elements containing ZpS1 satellite repeats have been investigated in a wide representation of species of the old plant genus Zamia (Zamiaceae, Cycadales). In Z. paucijuga, the ZpS1 repeat is organized as long satellite DNA arrays and as short arrays inserted into AT-rich dispersed elements. A comparative study by Southern analysis shows that these unusual dispersed elements containing the ZpS1 repeat are present with different organizations in all investigated Zamia species. In some species these elements are present with a low copy number, while in other species secondary amplification events, involving specific sequence clusters, appear to have generated characteristic dispersed elements in a high copy number. Among Zamia species, several groups share similar restriction patterns, as the Zamia loddigesii complex and the Caribbean species suggesting a general correlation between organization and genomic representation of the dispersed repeated sequence and the pattern of phyletic relationships in the genus. However, the finding of different patterns also among closely related species suggests a complex history of amplifications and losses of these dispersed repetitive elements that cannot be always easily traced through the phylogenetic reconstruction of this ancient plant group.     

The linear 20 kb mitochondrial genome of Pandorina morum (Volvocaceae,Chlorophyta)

A physical restriction map of the mitochondrial genome from one clone (TCC 854) of the sexually isolated populations (syngens) of the morphologically uniform species Pandorina morum Bory has been constructed using restriction endonucleases Ava I, Bam HI, Bgl II, Eco RI, Kpn I, and Pst I. The 20 kb linear genome can easily be separated from plastid DNA, nuclear satellite rDNA, and main band (nuclear) DNA on a Hoechst/CsCl buoyant density gradient. The Pandorina mitochondrial DNA shows sufficient similarity to the 16 kb mitochondrial genome of Chlamydomonas reinhardtii to cross-hybridize, and also hybridizes with a probe containing maize mitochondrial 18S rRNA genes. Double digests, self-probing, and Bal31 exonuclease experiments suggest that 1.8 to 3.3 kb of sequence is repeated at each end of the genome as an inverted repeat. Mitochondrial genome sizes of other P. morum syngens were found to range from ca. 20 to ca. 38 kb. The mitochondrial genome should be valuable for taxonomic studies; it can be used for comparative organellar studies; and it should be of interest to compare with that of other plant and animal mitochondrial genomes.     

A highly repeated DNA sequence in Arabidopsis thaliana

Summary Three members of a family of highly repeated DNA sequences from Arabidopsis thaliana have been cloned and characterized. The repeat unit has an average length of 180 bp and is tandemly repeated in arrays longer than 50 kb. This family represents more than one percent of the Arabidopsis genome. Sequence comparisons with tandemly repeated DNA sequences from other Cruciferae species show several regions of homology and a similar length of the repeat unit. Homologies are also found to highly repeated sequences from other plant species. When the sequence CCGG occurs in the repeated DNA, the inner cytosine is generally methylated.     

Molecular analysis of the genomic organization of the Hymenoptera Diadromus pulchellus and Eupelmus vuilleti

The genomic organization of two parasitic wasps was analyzed by DNA reassociation. Cot curves revealed a pattern with three types of components. A highly repetitive DNA, accounting for 15 to 25% of the genome, was identified as satellite DNA. The moderately repetitive DNA corresponds to 26 to 42% of the genome in both species, and shows large variations in complexity, repetitive frequency and a number of sub-components between males and females. These variations are seen as resulting from DNA amplification during somatic and sexual differentiation. Dot blot analyses show that such DNA amplifications concern several types of structural and regulatory genes. The presence of repeated mobile elements was studied by the Roninson method to compare the repeated sequence patterns of Diadromus pulchellus and Eupelmus vuilleti with those of Drosophila melanogaster. The occurrence and organization of mobile elements in these Hymenoptera differ from those of the neighboring order of Diptera. The repetitive and unique components define very large genomes (1 to 3 × 10⁹ base pairs). The genomic organization in Parasitica appears to be an extreme drosophilan type. We propose that the germinal genome of these parasitic wasps is primarily composed of satellite DNA blocks and very long stretches of unique sequences, separated by a few repeated and/or variously deleted, interspersed elements of each mobile element family.