Characterization of a new repetitive sequence that is enriched on microchromosomes of turkey

We cloned and characterized a new highly repetitive, species-specific DNA sequence from turkey (Meleagris gallopavo). This repeat family, which accounts for approximately 5% of the turkey genome, consists of a 41 bp repeated element that is present in tandem arrays longer than 23 kb. In situ hybridization to turkey metaphase chromosomes (2n=80) demonstrated that this sequence was located primarily on certain microchromosomes: approximately one-third of the 66 microchromosomes showed a positive signal. With respect to the macrochromosomes, hybridization was seen only in a pericentric position on nos. 2 and 3. The turkey microchromosome (TM) sequence shares motifs (alternating A_3–5 and T_3–5 clusters separated by 6–8 bp) that have been found previously in other avian tandemly repeated elements, e.g. a chicken microchromosome sequence, and W (female) chromosome-specific sequences of chicken and turkey. However, the TM sequence does not cross-hybridize under moderately stringent conditions with these other sequence. The spread and amplification of related repetitive sequence elements on microchromosomes and W chromosomes is discussed.by E.R. Schmidt     

The characterization of arabms1: an Arabidopsis thaliana sequence homologous to bacteriophage M13 repeat element

A genomic DNA fragment was isolated from the genome of Arabidopsisthaliana via hybridization with bacteriophage M13 protein IIIrepeat element. A 45 bp region of the A. thaliana DNA fragmenthas a repeating 12 bp structure that shows sequence homologyto both the M13 repeat and to a rice minisatellite-like element.Hybridization of digests of A. thaliana genomic DNA with theminisatellite DNA generates a multilocus DNA fingerprint withlow polymorphism. Key words: Minisatellite, M13 repeat element, Arabidopsis thaliana     

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.     

A novel avian W chromosome DNA repeat sequence in the lesser black-backed gull (Larus fuscus)

The phenol emulsion reassociation technique was used to isolate and clone a female specific, repetitive DNA sequence fromLarus fuscus. The repeat, designated P2000-17, is restricted to the W chromosome, although related sequences occur elsewhere in the genome ofL. fuscus. Similar sequences were detected in the genome of six other bird species from outside the genus Laridae, but the sequence occurs less frequently and to a similar extent in both sexes. The 298 bp DNA sequence of P2000-17 was determined and found to have extensive sequence identity to the rabbit dihydropyridine (DHP) receptor calcium channel. P2000-17 is represented once within a larger 8.6 kb tandem repeat (LfW-1), which has a complex internal DNA sequence. LfW-1 is highly conserved between repeat motifs and may comprise 3% of the female genome. The possible evolutionary origin of LfW-1 is discussed in relation to the repeat types found on the W and Y chromosomes of other species.     

A novel centromeric repetitive DNA from human chromosome 22

A recombinant DNA clone localized in the centromeric region of chromosome 22 was isolated from a flow-sorted human chromosome 22 DNA library. When the original insert of about 1.9 kb was used to probe Southern blots of EcoRI-digested genomic DNA it revealed at least 40 fragments. A comparable pattern was obtained with each of the three subclones (800, 700, and 380 bp). In situ hybridization showed signals clustered in the region 22cen. DNA sequence analysis using the 380 bp fragment subcloned in pTZ18/19 (p22hom48.4) revealed eight copies of a 48 bp repeat and the size of hybridizing restriction fragments indicated that this tandemly repeated sequence is spread over a region of a few hundred kilobases. Whereas this novel DNA, termed D22Z3, displayed no sequence homology to rodent and monkey genomes cross-homology was discernible for DNA from two great ape species.     

Characterization and localization of repetitive DNA sequences in the ornamental Alstroemeria aurea Graham

 Three repetitive DNA sequences were isolated from a genomic DNA library of the ornamental Alstroemeria aurea Graham. Two repeats, A001-I and A001-II, were quite homologous and highly A. aurea-specific. A001-I was a 217-bp sequence with several telomeric TTTAGGG repeats at the 5′ end and a unique sequence of 98 bp at the other end. The third repeat, A001-IV, was a 840-bp sequence which contained two sub-sequences of 56 and 74 bp respectively, previously found in chloroplast (cp) DNA of tobacco and spinach and to a lesser extent in the cpDNA of maize and rice. Repeat A001-IV was not species-specific and its hybridization signal was weaker than the other repeats. Fluorescence in situ hybridization (FISH) revealed the A. aurea-specific repeats to be located in the heterochromatic regions of all A. aurea chromosomes. The differences in FISH pattern make them useful tools for karyotype analysis. The non-species-specific sequence A001-IV gave a dispersed signal over all the Alstroemeria chromosomes in an interspecific hybrid. The potential use of these repetitive DNA sequences for the study of phylogenetic relationships within the genus Alstroemeria is discussed. Received: 24 November 1996/Accepted: 20 December 1996     

A Family of Differentially Amplified Repetitive DNA Sequences in the Genus Beta Reveals Genetic Variation in Beta vulgaris Subspecies and Cultivars

Members of a highly abundant restriction satellite family have been isolated from the wild beet species Beta nana. The satellite DNA sequence is characterized by a conserved RsaI restriction site and is present in three of four sections of the genus Beta, namely Nanae, Corollinae, and Beta. It was not detected in species of the evolutionary old section Procumbentes, suggesting its amplification after separation of this section. Sequences of eight monomers were aligned revealing a size variation from 209 to 233 bp and an AT content ranging from 56.5% to 60.5%. The similarity between monomers in B. nana varied from 77.7% to 92.2%. Diverged subfamilies were identified by sequence analysis and Southern hybridization. A comparative study of this repetitive DNA element by fluorescent in situ hybridization and Southern analyses in three representative species was performed showing a variable genomic organization and heterogeneous localizations along metaphase chromosomes both within and between species. In B. nana the copy number of this satellite, with some 30,000 per haploid genome, is more than tenfold higher than in Beta lomatogona and up to 200 times higher than in Beta vulgaris, indicating different levels of sequence amplification during evolution in the genus Beta. In sugar beet (B. vulgaris), the large-scale organization of this tandem repeat was examined by pulsed-field gel electrophoresis. Southern hybridization to genomic DNA digested with DraI demonstrated that satellite arrays are located in AT-rich regions and the tandem repeat is a useful probe for the detection of genetic variation in closely related B. vulgaris cultivars, accessions, and subspecies. Received: 24 May 1996 / Accepted: 13 September 1996     

Characterisation of a short interspersed repeat (Mermaid) that has family members on human chromosome 21 and elsewhere in the human genome

To understand the architecture of the human genome, we need a complete definition of all the repeat sequence families, as these make up the majority of human DNA. We have isolated a small DNA fragment from human chromosome 21 and have used sequence analysis of this fragment to uncover a new low copy repeat element of approximately 300 bp that we term the Mermaid repeat. This repeat is related to, but is different from, the MER 12 repeat and is interspersed in the genome. Mermaid family members that we have studied are between 81%–87% identical to our preliminary consensus sequence. Therefore, we have added a new member to the large collection of human repetitive elements. In addition, we have mapped a Mermaid repeat to a telomeric position on the long arm of human chromosome 21, at 21q22.3     

Origin and Differentiation of a Sex Chromosome System in Parodon hilarii (Pisces, Parodontidae). Satellite DNA, G- and C-banding

A satellite DNA sequence of Parodon hilarii (named pPh2004) was isolated, cloned and sequenced. This satellite DNA is composed of 200 bp, 60% AT rich. In situ hybridization (FISH) results revealed that the satellite DNA pPh2004 is located in the terminal regions of several chromosomes, forming highly evident blocks in some and punctual marks in others. The comparison between the FISH and C-banding results showed that the location of this satellite DNA coincides with that of most terminal heterochromatins. However, some regions are only marked by FISH whereas other regions are only marked by C-banding. The possible existence of more than one satellite DNA family could explain these partial differences. The in situ hybridization with the satellite DNA and the G- and C-bandings confirmed the presence of a sex chromosome system of the ZZ/ZW type in P. hilarii, as well as the correct identification of the Z chromosome in the karyotype. This chromosome displays a segment of terminal heterochromatin in the long arm, similar to the segment observed in the short arm of the W chromosome, also showing a G-banding pattern similar to that of the short arm and part of the long arm of the W chromosome. A hypothesis on the origin of the W chromosome from an ancestral chromosome similar to the Z chromosome is presented.     

Restriction enzyme analysis of the germ line limited DNA of Ascaris suum

The germ line limited DNA of Ascaris suum was isolated from sperm and testis as a satellite DNA component in Hoechst 33258 — CsCl gradients. Employing restriction enzyme analysis, we show that the germ line limited DNA is composed entirely of two families of tandemly repeated sequences, one repeat unit is 125 bp, and the other 131 bp long. The total appr. 5 × 10⁵ copies of the two families are physically separated from each other (segmental arrangement). Several repeat unit variants within both families could be detected. The copies of sequence variants are arranged in tandem (subsegmental arrangement). Reassociation and hybridization experiments revealed similar sequences of the two repeat units. The archaeotypic core sequence of both repeat units is probably a tetranucleotide which shows a theme and variation pattern. During chromatin diminution in the presoma cells the satellite DNA is eliminated from the chromosomes. However, a limited number of tandemly repeated copies of both kinds of repeat units could be detected in the soma genome using radioactive probes of both repeat units in Southern blots of muscle and intestine of adult animals. The tandem arrangement and the hierarchical pattern of restriction sites throughout different subfamilies supports the model of successive segmental amplification events during the evolution of the germ line limited DNA. Since the germ line limited satellite DNA is exclusively located at the ends of the chromosomes, a fold back structure for the telomeric DNA sequences is proposed which might have generated this DNA.     

Insertion and amplification of a DNA sequence in satellite DNA of Cucumis sativus L. (cucumber)

Summary Another satellite DNA repeat (type IV) in the genome of Cucumis sativus (cucumber) was found and investigated with respect to DNA sequence, methylation, and evolution. This satellite shows a repeat length of 360 bp and a GC-content of 47%. The repeats of type IV are highly conserved among each other. Evidence for CG and CNG methylation is presented. By comparison to the previously described satellites (type I/II and type III) from cucumber, it is evident that this repeat is created by an insertion of a 180 bp DNA sequence similar to type I–III into another DNA sequence (or vice versa), and subsequent amplification forming a new satellite repeat. The different satellites of the type I/II, type III, and the 180 bp insert of type IV show a sequence homology of 60%–70%, indicating that the complex satellite DNA of cucumber is originated from a common progenitor by mutation, additional insertion, and amplification events. Copies of a sequence similar to a part of type IV are present in the genome of the related species Cucumis melo (melon).     

A large DNA repeat of the dispersion pattern common to wheat and rye genomes

A Hind III-generated fragment of wheat embryo nuclear DNA has been cloned and sequenced. The cloned fragment corresponds to a 1241 bp long, moderately repeated (60 000 copies/genome) segment of the genomic DNA. The repeat is AT-rich (67%), contains an open reading frame for 151 amino acids and several nucleotide blocks resembling the consensus domain of autonomously replicating sequences. Southern blot hybridization analyses indicate that the repeat is scattered through the wheat genome. A sequence homologous to this repeat occurs also in rye embryo nuclear DNA where it shows the same dispersion pattern as that observed for the wheat repeat.     

A plasmid sequence from Rhizobium leguminosarum 300 contains homology to sequences near the octopine TL-DNA right border

Summary The DNA sequence from a Rhizobium leguminosarum 300 (RL300) plasmid that contains homology to the T_c-DNA of Agrobacterium tumefaciens is described. The RL300 sequence has 78% homology to a 359 bp sequence in the T_c-DNA of pTi15955. The RL300 homology starts approximately 100 bp from the 24 bp border sequence of the T_L-DNA and ends approximately 3 bp from an IS66 homolog in the T_c-DNA. An unusual feature of the RL300 homology is the presence of 81 bp direct repeats with T_c-DNA homology, separated by 201 bp. One end of each direct repeat has a 12 bp palindrome. Four cloned sequences of RL300 with homology to the T DNA region were hybridized to plasmid lysates of RL300 derivatives to determine the source of each plasmid. The sequenced homolog, originally on pRH228, was isolated from pRL7JI; the other 3 homologs were isolated from the transmissable plasmids pRL7JI (pRH235) and pRL8JI (pRH235 and pRH236).     

Molecular cloning of zebra finch W chromosome repetitive sequences: evolution of the avian W chromosome

The zebra finch (Taeniopygia guttata) has a large Z chromosome and highly condensed W chromosome. We used the random amplified polymorphic DNA (RAPD) polymerase chain reaction (PCR) technique to isolate female-specific sequences ZBM1 and ZBM2. Southern blot hybridization to male and female zebra finch genomic DNA suggested that these sequences were located on the W chromosome, although homologous sequences appeared to be autosomal or Z-linked. Fluorescent in situ hybridization (FISH) using bacterial artificial chromosome (BAC) clones corresponding to ZBM sequences showed hybridization to the whole W chromosome, suggesting that the BACs encode sequences that are repeated across the entire W chromosome. Based on the sequencing of a ZBM repetitive sequence and Z chromosome derived BAC clones, we demonstrate a random distribution of repeat sequences that are specific to the W chromosome or encoded by both Z and W. The positions of ZW-common repeat sequences mapped to a noncoding region of a Z chromosome BAC clone containing the CHD1Z gene. The apparent lineage-specificity of W chromosome repeat sequences in passerines and galliform birds suggest that the W chromosome had not differentiated well from the Z at the time of divergence of these lineages. Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

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.     

Organization of a Solatium brevidens repetitive sequence related to the TGRI subtelomeric repeats of Lycopersicon esculentum

A species-specific repetitive DNA fragment has been isolated from a genomic library of Solanum brevidens. Sequence analysis revealed a regular organization of three non-homologous subrepeats forming tandemly-arranged composite repetitive units. Interpretation of Southern hybridization patterns based on the known sequence data suggests that the isolated sequence element represents an abundant organization type, although the presence of simple tandem arrays of the subrepeats is also indicated. Seventy-four percent sequence similarity was found between one of the S. brevidens subrepeats (Sb4AX) and a satellite DNA (TGRI) localized as a subtelomeric repeat on almost all Lycopersicon esculentum chromosomes. Insitu hybridization indicated that, similarly to TGRI, the S. brevidens-specific repeats are located at the ends of the arms of several chromosomes. On the basis of the data obtained, a common ancestral sequence can be proposed for the tomato (TGRI) and the S. brevidens (Sb4AX) repeat however, the molecular organization of this element in these two species evolved in a basically different manner.     

Characterization of MboI repeat DNA sequence of Anopheles stephensi.

MboI repeat fragment of mosquito Anopheles stephensi has been isolated by molecular cloning. The restriction map and entire nucleotide sequence of the 433bp insert has been determined. Hybridization of this repeat DNA with restriction enzyme digest of mosquito DNA does not show an interspersed pattern but suggests that this repeat may be tandemly repeated at one major site and a few minor sites in the genome of Anopheles stephensi. The hybridization pattern also indicates that this repeat family comprises of many similar but non-identical sequences. An open reading frame encoding 66 amino acids with an initiation and two tandem termination codons has been identified. This putative 66 amino acid polypeptide sequence has significant homology to a small region of RNA tumour viral envelope protein.     

Characterization and chromosomal distribution of a novel satellite DNA sequence of Japanese quail (Coturnix coturnix japonica)

A novel satellite DNA sequence of Japanese quail (Coturnix coturnix japonica) was isolated from genomic DNA digested with restriction endonuclease, Bg/II. Sequence analysis of three different-size clones revealed the presence of a tandem array of a GC-rich 41 bp repeated element. This sequence was localized by fluorescence in situ hybridization (FISH) primarily to microchromosomes of Japanese quail (2n = 78); approximately 50 of the 66 microchromosomes showed positive signals, although hybridization signals were also detected on chromosomes 4 and W. This satellite DNA did not cross-hybridize with genomic DNA of chicken (Gallus gallus) and Chinese painted quail (Excalfactoria chinensis) under moderately stringent conditions, suggesting that this class of repetitive DNA sequences was species specific and fairly divergent in Galliformes species.     

Origin and molecular structure of a midget chromosome in a common wheat carrying rye cytoplasm

The origin and molecular structure of the midget chromosome that is retained in a common wheat with rye cytoplasm, were studied by using fluorescent in situ hybridization (FISH). FISH with biotinylated rye genomic DNA as a probe clearly showed that the midget chromosome had originated from certain part(s) of rye chromosome(s). The midget chromosome did not possess sequences similar to wheat rDNA nor to a rye telomeric sequence with a 350 bp repeat unit. However, another repetitive sequence (120 bp family) of rye was found to occur at one end of the midget chromosome. The telomeric repeat sequences from Arabidopsis thaliana cross-hybridized to both ends of the midget chromosome as well as to wheat chromosomes. From the results obtained in this and previous studies, it is assumed that the midget chromosome originated from part of a rye chromosome, most likely the centromeric region of chromosome 1R, and that the telomeric sequences were synthesized de novo.by R. Appels     

Isolation and analysis of a new hopper hAT transposon from the Bactrocera dorsalis white eye strain

A new hopper element belonging to the hAT transposon family was isolated from the white eye mutant strain of the Oriental fruit fly, Bactrocera dorsalis. Using the original hopper element sequence from the wild type Kahuku strain as a template, the new hopper was isolated by inverse and direct PCR. Nucleotide sequence analysis reveals a 3131 bp element with terminal and subterminal inverted repeat sequences, an 8 bp duplicated insertion site, and a conceptual translation yielding a single uninterrupted 650 amino acid open reading frame. The white eye hopper has structure more consistent with function than the Kahuku element, indicating that hopper is not an ancient relic. The hopper element remains distantly related to other known hAT elements including those from insects, and presently it is most similar to Activator-related elements discovered in the human genome. DNA hybridization studies indicate, however, that elements closely related to hopper exist in another bactrocerid species, the melonfly, B. cucurbitae.