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PubMed1984

Organization and expression of non-Alu family interspersed repetitive DNA sequences in the mouse genome.

The mouse genome is complex with regard to DNA sequence organization and transcriptional activity. To more fully understand the role of interspersed repetitive DNA sequences we have isolated and characterized five different mouse non-Alu DNA sequence families. We have found that: (1) the distribution of repetitive sequences is non-random in the genome; (2) two of the five families (Bam5 and R) were previously described by Fanning (1982) and Gebhard et al. (1982), respectively. We found that these two families are linked to each other and are found adjacent to seven of seven studied structural genes but in randomly selected DNA fragments showed much less significant linkage. (3) The position of the Bam5 and R family repeat units relative to beta-globin and relative to a housekeeping gene has been evolutionarily conserved in mice and humans. (4) Three previously undescribed families representing from 200 to 40,000 copies per genome have been characterized and shown to have equivalent human sequences. (5) All five families studied are represented in RNA polymerase II transcripts. Little RNA polymerase III transcription homologous to these three families could be detected. The structural and functional features of these five families defined in this paper provide a basis for studies on the functional role of interspersed repetitive DNA in the mouse.

Animals↗
PubMed1995

Use of interspersed repetitive sequences-PCR products for cDNA selection.

In order to increase the efficiency of cDNA selection approaches, we describe the use of interspersed repetitive sequences-PCR (IRS-PCR) products to isolate genes from large-insert genomic clones. IRS-PCR is conducted on total yeast DNA containing a YAC of interest so that there is no need to purify the starting genomic clone. This enables the production of large amounts of genomic substrate for cDNA selection and allows the use of unstable YAC clones. Moreover, the hybridization of the IRS-PCR product to the cDNA clones after selection introduces a positive selection step. We tested these PCR products from YACs for the presence of exons, using cDNAs originating from seven different genes. In each case, at least one exon was present in the IRS-PCR product. We have applied this strategy to four YAC clones originating from the human X Chromosome (Chr). All the selected cDNAs, strongly positive with the IRS-PCR product, did indeed originate from a gene in the region covered by the YAC. In all cases, the previously known genes contained in the genomic clones have been isolated. In addition, we have isolated human genes that have already been described but not assigned to any chromosomal region.

Base Sequence↗
PubMed1984

Clustered and interspersed repetitive DNA sequence family of Chironomus. The nucleotide sequence of the Cla-elements and of various flanking sequences.

The nucleotide sequence of more than 30 cloned members of the clustered and interspersed repetitive Cla-sequence family present in the genome of various chironomids has been determined. In four cloned Cla-element clusters, the 5' and 3'-flanking sequences including the junctions between the Cla-element clusters and the flanking sequences were also sequenced. The repetitive Cla-elements, which are able to transpose under certain circumstances, have a monomer length ranging from 110 to 119 base-pairs, are very A + T-rich (greater than 80% A + T) and display numerous palindromic sequences. The Cla-elements are organized in small (4 elements) to medium-sized (greater than 30 elements) tandem repetitive clusters, which are dispersed over more than 200 sites of the chromosomes of Chironomus thummi thummi, including the non-transcribed spacer of the ribosomal DNA repeating unit. The tandem repetitive Cla-elements show anomalous behaviour during high-percentage polyacrylamide gel electrophoresis, indicating a bent or globular conformation. The flanking sequences are also repetitive, but the sequenced parts did not reveal any tandem repetitive arrangement. Near the junctions of the Cla-element clusters and the flanking sequences, short duplications are found, ranging from 5 to 12 bases, present in both sides of the Cla-element clusters. The Cla-elements might be involved in the hybrid dysgenesis phenomenon that is observed after crossings between the two subspecies Ch. th. thummi and Ch. th. piger.

Animals↗
PubMedDate not supplied

Transcription of the KpnI families of long interspersed DNAs in human cells.

The mammalian genome contains a variety of interspersed repetitive sequences of unknown function. It has, however, been suggested that interspersed repetitive sequences and their RNA transcripts are involved in the coordinate regulation of gene expression. Two major families of interspersed sequences in primates are the so-called Alu and KpnI families. Members of the KpnI families range in length from 1.2 to over 6 kilobases (kb). They exist in generally clustered arrangements, in 6 X 10(4) to 10(5) copies per diploid genome. Something is known of the arrangements of KpnI family sequences near human structural genes, but there has been no information on transcription of the sequences. We report here that the KpnI sequences are transcribed in HeLa cells by RNA polymerase II into abundant and heterogeneous species of RNA. The transcripts range in length from about 200 bases to over 5 kb, and are found predominantly in the non-polyadenylated fraction of the nuclear RNA. Transcripts homologous to both of the complementary strands of the KpnI sequences are present, but with a strong bias towards one strand.

Base Sequence↗
PubMed1992

Integration of bovine papillomavirus type 1 DNA and analysis of the amplified virus-cell junctions in transformed primary mouse fibroblasts.

We have analysed the site of bovine papillomavirus type 1 (BPV-1) DNA integration in clones originating from a transformed primary mouse fibroblast cell line established by transfection of linear BPV-1 DNA. Viral DNA was integrated at a single site in the host genome with an intact early region and an almost complete long control region. Sequence analysis showed that the BPV-1 DNA was integrated at the HindIII site (the enzyme used to linearize the BPV-1 DNA for transfection) with short deletions at both ends. These deletions correspond to a 534 bp segment spanning the 3' end of the L1 open reading frame and the replication enhancer element in the BPV-1 genome. The cellular sequences 5' to the viral integration site exhibited 85 to 97% identity to several sequences belonging to the mouse L1 family of long interspersed repetitive sequences. Cellular sequences 3' to the viral DNA exhibited no significant similarity to any known sequence. The BPV-1 sequences and the cellular flanking sequences were found to be amplified 45- to 50-fold. All the cell clones shared an identical integration site but one of the clones had an additional population of amplified and integrated BPV-1 DNA molecules with an internal deletion of 1136 bp in the late region. The significance of viral DNA integration at a murine long interspersed repetitive sequence containing an amplification-promoting sequence is discussed.

Animals↗
PubMed1986

Replication time of interspersed repetitive DNA sequences in hamsters.

The replication time of 34 hamster genomic DNA segments containing interspersed repeat sequences was determined by probing the cloned segments with nick-translated early- and late-replicating hamster DNA. One-third of these cloned families replicated early, one-third replicated late, and one-third replicated without temporal bias. 19 different inserts from these clones along with the SINE, Alu, and the LINE, A36Fc, were used to probe Southern blots of early- and late-replicating hamster or human DNA. We report long interspersed repeats, LINEs, are selectively partitioned into late-replicating DNA and are often concertedly hypomethylated, while short interspersed repeats, SINEs, are selectively partitioned into early-replicating DNA. For some interspersed repeat families, this partitioning is complete or almost complete. The CCGG frequency is very low in late-replicating DNA. The mammalian chromosome's pattern of early-replicating R-bands and late-replicating G-bands reflects a differential distribution of LINEs and SINEs.

Animals↗
PubMed1992

Rapid epidemiologic analysis of cytomegalovirus by using polymerase chain reaction amplification of the L-S junction region.

A technique based on polymerase chain reaction (PCR) amplification was developed to facilitate the study of the epidemiology of cytomegalovirus (CMV). Consensus oligonucleotide primers from repetitive DNA sequences were designed to amplify interspersed repetitive sequences in an area of heterogeneity within the L-S junction region of the CMV genome, and PCR products were detected by gel electrophoresis. Purified CMV DNAs from 25 CMV isolates, 13 from members of five families in which person-to-person transmission was documented, 9 random clinical isolates of CMV, and 3 laboratory reference strains of CMV (Towne, Davis, and AD169), were analyzed. The gel electrophoretic patterns of DNA bands, or PCR profiles, produced by amplification with the L-S primers were unique for epidemiologically unrelated strains and laboratory reference strains, yet similar patterns were observed for epidemiologically related strains isolated from members of the same family. This method of rapid fingerprinting of CMV DNA within the hypervariable L-S junction region by PCR to produce strain-specific, variably sized PCR products should simplify the molecular epidemiologic analysis of CMV.

Base Sequence↗
PubMed1977

Length and interspersion of repetitive and non repetitive DNA sequences in four amphibian species with different genome sizes.

The interspersion period of repetitive and unique sequences was analyzed by two different methods, electron microscopy and agarose gel electrophoresis, for four Amphibian species with different nuclear DNA content, namely the Anura Xenopus laevis (3 pg DNA per haploid genome) and Bufo bufo (7 pg) and the Urodela Triturus cristatus (23 pg) and Necturus maculosus (52 pg). Within each of the two subclasses it has been found that interspecific differences, in DNA content, due to variations in the amount of repetitive sequences, do not involve variations in length of the interspersed repetitive sequences. They remain about 380 base pairs. Furthermore, the unique sequences length has been found to be shorter in Bufo (760 base pairs) than in Xenopus (1600) and in Necturus (880) than in Triturus (1340). A study of the interspersion period has shown that the great difference in DNA content between Anura and Urodela, which had been previously shown not to have involved changes in the relative amounts of the various sequence classes, does not involve changes in the interspersion period.

Animals↗
PubMed1990

Alu interspersed repeats: selfish DNA or a functional gene family?

Within the genomes of higher eukaryotic cells, short interspersed repetitive sequences appear to be ubiquitous, but also remarkably varied with respect to copy number and position. Many of these repeat families, including the human Alu family, can be transcribed by RNA polymerase III, and evidence has accumulated from a variety of sources that levels of repeat transcripts whose transcription is dependent on RNA polymerase III are sensitive to cellular transformation as well as changes in differentiation state. Although interspersed repetitive sequences have in the past been dismissed as nonfunctional, the discovery of the linkage to differentiation state, as well as other recent developments, suggest that the question of repeat sequence functionality should be reexamined.

Animals↗
PubMed1983

Total DNA transcription reveals the existence of highly repetitive transcribable sequences in higher animals.

Using HeLa S-100 extracts, we have transcribed total DNA isolated from several species of higher animals. Transcription of the total DNA from rat, mouse, or hamster gives a discrete product of 6S RNA (about 180 nucleotides) in addition to a smeared background, whereas transcription from chick, calf, or human DNA gives a discrete band of 5S RNA (about 120 nucleotides). In view of the sensitivity to a-amanitin, these transcripts are produced by RNA polymerase III. Fingerprint analysis demonstrated that the 6S RNA species transcribed from rodent DNA have conserved sequences. The composition of dinucleotides in these bands having a guanosine residue on the 3' side was analyzed by RNase T2 digestion. The results show that the CG ratios of the transcripts of rodent, calf and chick are quite low in comparison with AG, UG, or GG except in the case of the human transcript, suggesting that these highly repetitive transcribable sequences are derived from a common prototype sequence, except in human cells. We have isolated two phage clones and one phage clone from a rat and mouse genomic library, respectively, which give a transcript of 180 nucleotides in size, and whose pattern of RNase T2 digests is the same as that of the conserved transcriptional products of total DNA. These highly repetitive transcribable sequences appear to be reiterated at several to ten thousand copies per haploid genome. The relationship of these sequences with the highly repetitive interspersed sequence B1 or Alu family is discussed.

Animals↗
PubMed1983

Organization of the R family and other interspersed repetitive DNA sequences in the mouse genome.

Six R sequences have been described as members of a new family of dispersed repetitive DNA in the mouse genome (Gebhard et al., 1982). Several sequenced regions were extended in the 5' direction (R1, R2 and R6) and two new sequences were determined (R7 and R8). On the basis of our sequence and blot hybridization data it is concluded that the R sequence are adjacent to the so-called small Bam family (Fanning, 1982), which in turn runs into the MIF sequence part (Brown & Piechaczyk, 1983) of the large Bam sequences (Meunier-Rotival et al., 1982). In one of our clones a sequence of 1290 base-pairs comprises MIF, Bam and R sequences in a contiguous arrangement which seems to be characteristic of the long repeat unit of the mouse genome. Several repeat units were found to be truncated within their Bam or R sequence parts. Evidence is also reported for transposition events involving R sequences; for instance of one R sequence (R1) into another (R7). Two R sequences (R1 and R4) have apparently been transposed together with part of the adjacent Bam sequences. Truncation and transposition events may also explain the imbalance of copy numbers within the large repeat unit (25,000 to 50,000 for the small Bam sequences and 100,000 for the R sequences). The spreading of R sequences and other interspersed DNA sequences within the mouse genome may have occurred by transposition events on the DNA level and/or by transcription, retrotranscription and insertion processes.

Animals↗
PubMed1987

Structure and evolution of a family of interspersed repetitive DNA sequences in Caenorhabditis elegans.

The structure of three members of a repetitive DNA family from the genome of the nematode Caenorhabditis elegans has been studied. The three repetitive elements have a similar unitary structure consisting of two 451-bp sequences in inverted orientation separated by 491 bp, 1.5 kb, and 2.5 kb, respectively. The 491-bp sequence separating the inverted 451-bp sequences of the shortest element is found adjacent to one of the repeats in the other two elements as well. The combination of the three sequences we define as the basic repetitive unit. Comparison of the nucleotide sequences of the three elements has allowed the identification of the one most closely resembling the primordial repetitive element. Additionally, a process of co-evolution is evident that results in the introduction of identical sequence changes into both copies of the inverted sequence within a single unit. Possible mechanisms are discussed for the homogenization of these sequences. A direct test of one possible homogenization mechanism, namely homologous recombination between the inverted sequences accompanied by gene conversion, shows that recombination between the inverted repeats does not occur at high frequency.

Animals↗
PubMed1984

Dispersal process associated with the L1 family of interspersed repetitive DNA sequences.

We have determined the complete nucleotide sequence for five members of the L1Md repetitive family from the beta-globin gene region of the BALB/c mouse. The five repeats are different lengths, each terminating at the 5' end at different points with respect to one another. We have analyzed the nucleotides around the endpoints of the five repeats for clues as to the mechanisms involved with the dispersal and 5' truncation of this repeat family. Each L1 member is flanked by a pair of short direct repeats. Since these direct repeats differ in length and sequence in each of the five cases, the dispersal mechanism does not involve a sequence targeted process. The sequence at the 3' end is conserved and its organization resembles the 3' end of a polyadenylated RNA, suggesting that transcripts of the repeat are involved in the dispersal process either directly or as intermediates in the generation of complementary DNA copies of the sequence. One of the L1 repeats is a recent insertion, since it is found in the Hbbd chromosome, but not in the Hbbs chromosome. This suggests a dispersal process that has been active as recently as 4 million years ago.

Animals↗