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Results for “DNA, Circular”

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At least 19 recordsLinked to original sources
PubMed1980

Homologous pairing in genetic recombination: recA protein makes joint molecules of gapped circular DNA and closed circular DNA.

The recA protein, which is essential for genetic recombination in E. coli, promotes the homologous pairing of double-stranded DNA and linear single-stranded DNA, thereby forming a three-stranded joint molecule called a D loop. Single-stranded DNA stimulates recA protein to unwind double-stranded DNA. By a presumably related mechanism, recA protein promoted the homologous pairing of two circular double-stranded molecules when one of them has a gap in one strand. The two molecules were joined at homologous sites by noncovalent bonds. The covalently closed molecule remained intact and was not topologically linked to the intact circular strand of the gapped substrate. Electron microscopy showed that molecules were usually linked at two or more nearby points. The junctions in most molecules were shorter than 300 nucleotides. Sometimes the region between two extreme points was separated into two arms, producing an ellipsoidal loop (called an eye loop). The junctions in these biparental joint molecules were frequently remote from the site of the gap. We infer that a free end of the interrupted strand crosseover to form a structure like a D loop which moved away from the gap by branch migration.

Bacterial Proteins↗
PubMed2004

Somatic DNA recombination yielding circular DNA and deletion of a genomic region in embryonic brain.

In this study, a mouse genomic region is identified that undergoes DNA rearrangement and yields circular DNA in brain during embryogenesis. External region-directed inverse polymerase chain reaction on circular DNA extracted from late embryonic brain tissue repeatedly detected DNA of this region containing recombination joints. Wide-range genomic PCR and digestion-circularization PCR analysis showed this region underwent recombination accompanied with deletion of intervening sequences, including the circularized regions. This region was mapped by fluorescence in situ hybridization to C1 on mouse chromosome 16, where no gene and no physiological DNA rearrangement had been identified. DNA sequence in the region has segmental homology to an orthologous region on human chromosome 3q.13. These observations demonstrated somatic DNA recombination yielding genomic deletions in brain during embryogenesis.

Animals↗
PubMed1980

Targeted deletions of sequences from closed circular DNA.

Closed circular DNA interacts with complementary sequences of single-stranded DNA to form displacement loop (D loop) structures in vitro. The site of D-loop formation can be directed by using single-stranded DNA derived from a selected restriction fragment. Circular DNA containing a D loop can then be linearized by cleavage with endonuclease S1. This cleavage appears to remove a limited number of nucleotides from each strand of the circular DNA substrate. Incubation with polynucleotide ligase followed by propagation in vivo leads to circular DNA molecules that bear small, single deletions in the region of the single-stranded DNA sequence chosen for the formation of the D loops. We have utilized these manipulations of DNA to construct tetracycline-sensitive deletion mutants of plasmid pBR322. The level of mutagenesis obtained by the procedure is sufficiently high that selective growth and screening procedures are not necessary for the isolation or identification of mutants. The frequency, variety, and small size of the deletions obtained within the selected target regions present considerable advantage for genetic and biochemical analysis. The method is quite general in rationale and should be immediately applicable to phage and viruses having infectious circular DNA genomes or recombinant DNA species propagated in circular plasmid vectors.

Chromosome Deletion↗
PubMed1985

L1 sequences in HeLa extrachromosomal circular DNA: evidence for circularization by homologous recombination.

Subcloned probes of the L1 family of repetitive elements were used to isolate L1-carrying clones from a plasmid library of HeLa cell extrachromosomal circular DNA. One clone was analyzed in detail by restriction mapping, cross-hybridization to L1 probes, and base sequence analysis. In addition to approximately the 3' half of a full-sized L1 element, this clone carried 390 base pairs of non-L1 sequence that is single copy in the HeLa genome. A HeLa genomic clone of this unique chromosomal region was isolated and the sequence organization of the circle clone was compared with the linear chromosomal region from which it was ultimately derived. We discuss possible mechanisms of circular DNA formation and propose homologous intrachromosomal recombination between 9-base-pair direct repeats to be most likely in this case.

Chromosome Deletion↗
PubMed1979

Mitochondrial DNA from Podospora anserina. II. Properties of mutant DNA and multimeric circular DNA from senescent cultures.

Mitochondrial (Mt) DNA from mitochondrial mutants of race s Podospora anserina and from senescent cultures of races s and A was examined. In mutants, we observed that fewer full length circles (31 mu) were present; instead, smaller circles characteristic for each mutant studied were found. Eco R1 digestion of these mutant MtDNAs indicated that in certain mutants, although specific fragments were absent, the total molecular weight of the fragments was not much different than wild-type. The properties of senescent MtDNA was strikingly different from either wild-type or mutant Mt DNA. First, a multimeric set of circular DNA was observed for both race s and A, with a monomeric repeat size of 0.89 mu. These circles ranged in size from 0.89 mu to greater than 20 mu; only one molecule out of some 200 molecules was thought to be of full length (31 mu). Density gradient analysis showed that there were two density species: a majority were at the same density as wild-type (1.694 g/cm3) and a second at 1.699 g/cm3. Most of the circular molecules from MtDNA isolated by either total DNA extraction or by extraction of DNA from isolated mitochondria were contained in the heavy DNA fraction. Eco R1 enzymatic digestion indicated that the light DNA had several fragments (amounting to about 23 x 10(6) daltons) missing, compared with young, wild-type MtDNA. Heavy senescent MtDNA was not cleaved by Eco R1. Analysis with Hae III restriction endonuclease showed also that light senescent MtDNA was missing certain fragments. Heavy MtDNA of average size 20 x 10(6) daltons, yielded only one fragment, 2,500 bp long, by digestion with Hae III restriction endonuclease. Digestion of heavy DNA with Alu I enzyme yielded 10 fragments totalling 2,570 bp. By three criteria, electron-microscopy, Eco R1 and Hae digestion, we conclude that the heavy MtDNA isolated from senescent cultures of Podospora anserina consisted of a monomeric tandemly repeating subunit of about 2,600 bp length. These results on the properties of senescent MtDNA are discussed with regard to the published properties of the rho- mutation in the yeast, S. cerevisiae.

Ascomycota↗
PubMed2025

eccDNABase: A Comprehensive and High-Quality Database for Extrachromosomal Circular DNA.

Extrachromosomal circular DNA (eccDNA) refers to small, circular DNA molecules that originate from chromosomal sequences and are prevalent across nearly all eukaryotic organisms. In humans, eccDNAs are widely distributed in normal tissues, cancerous tissues, and body fluids, where they play important roles in tumorigenesis and are often associated with poor clinical outcomes. Given their biological and clinical significance, a well-integrated and high-quality database is essential for advancing eccDNA-related research. To address this need, we developed eccDNABase, a comprehensive and curated resource for browsing, searching, and analyzing eccDNAs across multiple species. The database systematically catalogs eccDNA-disease associations from diverse tissues and organisms. Currently, eccDNABase contains 1,875,452 eccDNA-disease associations, encompassing 8,398 ecDNA entries across nine species, 63 diseases, and healthy individuals. Each entry provides detailed information, including eccDNA ID, type, chromosomal localization, species, tissue or cell line source, disease name and Disease Ontology ID, overlap length and percentage with genes, oncogene overlap, detection method, and links to literature and source databases. Given its extensive and curated datasets, eccDNABase serves as a valuable resource for both basic and translational research, offering deeper insights into the role of eccDNA in health and disease. The database is publicly accessible at http://cgga.org.cn/eccDNABase/.

Humans↗
PubMed1992

Amplicon structure in multidrug-resistant murine cells: a nonrearranged region of genomic DNA corresponding to large circular DNA.

Multidrug resistance (MDR) in tumor cell lines is frequently correlated with amplification of one or more mdr genes. Usually the amplified domain also includes several neighboring genes. Using pulsed-field gel electrophoresis, we have established a restriction map covering approximately 2,200 kb in the drug-sensitive mouse tumor cell line TC13K. The mapped region is located on mouse chromosome 5 and includes the three mdr genes, the gene for the calcium-binding sorcin protein, and a gene with unknown function designated class 5. Long-range maps of the amplified DNA sequences in five of six MDR sublines that had been independently derived from TC13K generally displayed the same pattern as did the parental cell line. All six MDR sublines exhibited numerous double minutes, and one of them displayed a homogeneously staining region in a subpopulation. Large circular molecules, most likely identical to one chromatid of the double minutes, were detected in four of the sublines by linearization with gamma irradiation. The size of the circles was about 2,500 kb, which correlated to a single unit of the amplified domain. We therefore propose that in four independent instances of MDR development, a single unit of about 2,500 kb has been amplified in the form of circular DNA molecules. The restriction enzyme map of the amplified unit is unchanged compared with that of the parental cell line, whereas the joining sites of the circular DNA molecules are not identical but are in the same region.

Animals↗
PubMed1976

Restriction assay for integrative recombination of bacteriophage lambda DNA in vitro: requirement for closed circular DNA substrate.

A novel assay has been developed for in vitro genetic recombination of DNA. Substrate and product DNAs are cleaved with a restriction endonuclease and the resulting fragments are separated by electrophoresis in agarose gels. The substrate DNA has been chosen so that the recombination to be studied deletes a segment of DNA. The remaining DNA gives rise to a unique restriciton fragment, as does the DNA segment that has been removed. The method provides a convenient and physical, rather than genetic, assessment of the conversion of parental to recombinant DNA. This method has been applied to an in vitro system that carries out integrative recombination of bacteriophage lambda. We find that, different molecular forms of DNA tested, closed circular DNA is the only efficient substrate. Linear DNA and three kinds of circular DNA containing interruptions are at best very poor substrates. The implications of this surprising result are discussed. In addition, we show that the in vitro recombination system completes the breaking and rejoining steps of recombination. No stable DNA intermediates involving chiasmata or broken end structures are found.

Chromosome Mapping↗
PubMed1990

Circular DNA is a product of the immunoglobulin class switch rearrangement.

The class of immunoglobulin is defined by the constant region of its heavy chain. When a B lymphocyte switches the class of heavy chain it produces, the constant region of mu-type heavy chain is replaced; this occurs through a DNA rearrangement that brings the gene segment encoding the new constant region close to the VDJ segment encoding the variable region. The pre-B-cell line 18-81, which switches from heavy chain mu to gamma 2b production in culture, occasionally abnormally rearranges the heavy chain locus so that DNA sequences between the switch regions of mu and gamma 2b are inverted. Because looping-out is an intermediate step in generating an inversion, the switch rearrangement could occur by looping-out and deletion. Provided that recombination is reciprocal, this would produce a circle of DNA. Indeed, circular DNA molecules have been isolated as products of rearrangement among gene segments encoding the variable regions of the T-cell receptor and of the immunoglobulin heavy chain and light chain. But whereas the breakpoints for the variable region rearrangement are precisely defined, the breakpoints for any given heavy chain class switch are scattered over a length of greater than 6 kilobases, including both switch regions. We have now isolated circular DNA containing the sequences deleted by class-switching, thereby showing that the immunoglobulin heavy chain class switch occurs through looping-out and deletion.

Animals↗
PubMed2006

Real-time PCR quantitation of hepatitis B virus total DNA and covalently closed circular DNA in peripheral blood mononuclear cells from hepatitis B virus-infected patients.

It remains unclear whether hepatitis B virus (HBV) replicates in extrahepatic tissues, and particularly in peripheral blood mononuclear cells (PBMCs), which may serve as a reservoir for the maintenance of infection. A real-time PCR assay for the quantitation of total and covalently closed circular (ccc) HBV DNA in serum and in PBMCs was developed. This assay was highly sensitive (detection limit: 27 IU/mL), linear over a wide range (9 log10), and was displayed high inter- and intra-assay reproducibility for the quantitation of total DNA. Genotypes A to E were detected and the results were consistent with those obtained with the COBAS Amplicor HBV Monitor Test. The specificity of the methodology was increased by prior treatment with an enzyme that digests relaxed circular DNA, and the elimination of background signals from virus adsorbed to the surface of PBMCs. HBV DNA was detected in the serum and PBMCs of 12 HBsAg-positive patients, with less than 1% in the cccDNA form. In conclusion, the quantitation of total and ccc HBV DNA in PBMCs is potentially useful as a non-invasive marker, and may help to increase our knowledge of the natural history of hepatitis B.

DNA, Circular↗
PubMed2002

Inhibitory effect of adefovir on viral DNA synthesis and covalently closed circular DNA formation in duck hepatitis B virus-infected hepatocytes in vivo and in vitro.

The elimination of viral covalently closed circular DNA (CCC DNA) from the nucleus of infected hepatocytes is an obstacle to achieving sustained viral clearance during antiviral therapy of chronic hepatitis B virus (HBV) infection. The aim of our study was to determine whether treatment with adefovir, a new acyclic nucleoside phosphonate, the prodrug of which, adefovir dipivoxil, is in clinical evaluation, is able to suppress viral CCC DNA both in vitro and in vivo using the duck HBV (DHBV) model. First, the effect of adefovir on viral CCC DNA synthesis was examined with primary cultures of DHBV-infected fetal hepatocytes. Adefovir was administered for six consecutive days starting one day before or four days after DHBV inoculation. Dose-dependent inhibition of both virion release in culture supernatants and synthesis of intracellular viral DNA was observed. Although CCC DNA amplification was inhibited by adefovir, CCC DNA was not eliminated by antiviral treatment and the de novo formation of CCC DNA was not prevented by pretreatment of the cells. Next, preventive treatment of experimentally infected ducklings with lamivudine or adefovir revealed that both efficiently suppressed viremia and intrahepatic DNA. However, persistence of viral DNA even when detectable only by PCR was associated with a recurrence of viral replication following drug withdrawal. Taken together, our results demonstrate that adefovir is a potent inhibitor of DHBV replication that inhibits CCC DNA amplification but does not effectively prevent the formation of CCC DNA from incoming viral genomes.

Adenine↗