scholarly journals Clinical Validation of Fetal cfDNA Analysis Using Rolling-Circle-Replication and Imaging Technology in Osaka (CRITO Study)

Diagnostics ◽  
2021 ◽  
Vol 11 (10) ◽  
pp. 1837
Author(s):  
Ritsuko Kimata Pooh ◽  
Chika Masuda ◽  
Risa Matsushika ◽  
Megumi Machida ◽  
Takako Nakamura ◽  
...  
Keyword(s):  
Genetic Research ◽  
Rolling Circle Replication ◽  
Fetal Ultrasound ◽  
Aneuploidy Screening ◽  
Rolling Circle ◽  
Invasive Test ◽  
First Choice ◽  
Ultrasound Findings ◽  
Potential Applications ◽  
Monosomy 21

Background: Noninvasive prenatal genetic testing (NIPT) has been adopted as the first choice for aneuploidy screening. The purposes of this study were to investigate the accuracy of Vanadis® NIPT (hereafter CRITO-NIPT) in order to gain a deeper insight into the reasons for discrepancies, as well as to discuss the role of fetal ultrasound. Methods: Between 2019 and 2020, CRITO-NIPT was performed in 1218 cases of patients who underwent CVS or amniocentesis after a detailed fetal ultrasound exam and genetic counseling. The CRITO-NIPT results were compared with the genetic results. In cases of test discrepancies, the placentae were collected for detailed genetic research, and the pre-procedure fetal ultrasound findings were referred to. Results: The positive predictive value of T21, T18, and T13 was 93.55%, 88.46%, and 100%, respectively. In 90% of the of false positive (FP) cases, the placentae were examined. In 75% of the CRITO FP-T21 cases, placental mosaicism, or a demised twin’s T21, were confirmed. There were complicated mosaic cases, including tetrasomy 21/trisomy7 and monosomy 21/trisomy21 cases. In one of three no-call cases, an intermediate deletion of chromosome 13 was detected. Conclusions: The CRITO study investigated the mechanism of false positives, and the detailed mechanisms in mosaic and no-call cases. There have hitherto been no reports that have provided insight by partitioning the placenta to compare the NIPT and invasive test results, nor that have provided detailed ultrasound findings in the cases of discordant results, revealing the demonstrated importance of, and necessity for, detailed ultrasonography. This article describes the potential of rolling-circle replication as a powerful biosensing platform, as well as the importance of examining the fetus in detail with ultrasound. However, we should remember that the potential applications raise ethical and social concerns that go beyond aneuploidy and its methodology.

scholarly journals Targeted Allele Replacement Mutagenesis of Corynebacterium pseudotuberculosis

2011 ◽  
Vol 77 (10) ◽  
pp. 3532-3535 ◽  
Author(s):  
Caray A. Walker ◽  
Willie Donachie ◽  
David G. E. Smith ◽  
Michael C. Fontaine
Keyword(s):  
Corynebacterium Pseudotuberculosis ◽  
Sequence Length ◽  
Homologous Sequence ◽  
Proof Of Concept ◽  
Rolling Circle Replication ◽  
Content Type ◽  
Rolling Circle ◽  
Allele Replacement ◽  
Marker Free ◽  
The Relationship

ABSTRACTA two-step allele replacement mutagenesis procedure, using a conditionally replicating plasmid, was developed to allow the creation of targeted, marker-free mutations inCorynebacterium pseudotuberculosis. The relationship between homologous sequence length and recombination frequency was determined, and enhanced plasmid excision was observed due to the rolling-circle replication of the mutagenesis vector. Furthermore, an antibiotic enrichment procedure was applied to improve the recovery of mutants. Subsequently, as proof of concept, a marker-free,cp40-deficient mutant ofC. pseudotuberculosiswas constructed.

Homogeneous Detection of Single Rolling Circle Replication Products

2004 ◽  
Vol 76 (2) ◽  
pp. 495-498 ◽  
Author(s):  
Gerhard A. Blab ◽  
Thomas Schmidt ◽  
Mats Nilsson
Keyword(s):  
Rolling Circle Replication ◽  
Rolling Circle

scholarly journals Pf Filamentous Phage Requires UvrD for Replication in Pseudomonas aeruginosa

mSphere ◽  
2016 ◽  
Vol 1 (1) ◽  
Author(s):  
Eriel Martínez ◽  
Javier Campos-Gómez
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Dna Repair ◽  
Dna Helicase ◽  
Filamentous Phage ◽  
Rolling Circle Replication ◽  
Content Type ◽  
Rolling Circle ◽  
Initiator Protein ◽  
Biofilm Development ◽  
Filamentous Phages

ABSTRACT Biofilm development is a key component of the ability of Pseudomonas aeruginosa to evade host immune defenses and resist multiple drugs. Induction of the filamentous phage Pf, which usually is lysogenized in clinical and environmental isolates of P. aeruginosa, plays an important role in biofilm assembly, maturation, and dispersal. Despite the clinical relevance of Pf, the molecular biology of this phage is largely unknown. In this study, we found that rolling circle replication of Pf depends on UvrD, a DNA helicase normally involved in DNA repair. We also identified the initiator protein of Pf and found that it shares structural similarity with that of Vibrio cholerae phages CTXφ and VGJφ, which also use UvrD for replication. Our results reveal that, in addition to DNA repair, UvrD plays an essential role in rolling circle replication of filamentous phages among diverse bacteria genera, adding a new, previously unrecognized function of this accessory helicase. Pf is a lysogenic filamentous phage that promotes biofilm development in Pseudomonas aeruginosa. Pf replicates by a rolling circle replication system which depends on a phage-encoded initiator protein and host factors usually involved in chromosome replication. Rep, an accessory replicative DNA helicase, is crucial for replication of filamentous phages in Escherichia coli. In contrast, here we show that, instead of depending on Rep, Pf replication depends on UvrD, an accessory helicase implicated in DNA repair. In this study, we also identified the initiator protein of Pf and found that it shares similarities with that of Vibrio phages CTXφ and VGJφ, which also depend on UvrD for replication. A structural comparative analysis of the initiator proteins of most known filamentous phages described thus far suggested that UvrD, known as a nonreplicative helicase, is involved in rolling circle replication of filamentous phages in diverse bacteria genera. This report consolidates knowledge on the new role of UvrD in filamentous phage replication, a function previously thought to be exclusive of Rep helicase. IMPORTANCE Biofilm development is a key component of the ability of Pseudomonas aeruginosa to evade host immune defenses and resist multiple drugs. Induction of the filamentous phage Pf, which usually is lysogenized in clinical and environmental isolates of P. aeruginosa, plays an important role in biofilm assembly, maturation, and dispersal. Despite the clinical relevance of Pf, the molecular biology of this phage is largely unknown. In this study, we found that rolling circle replication of Pf depends on UvrD, a DNA helicase normally involved in DNA repair. We also identified the initiator protein of Pf and found that it shares structural similarity with that of Vibrio cholerae phages CTXφ and VGJφ, which also use UvrD for replication. Our results reveal that, in addition to DNA repair, UvrD plays an essential role in rolling circle replication of filamentous phages among diverse bacteria genera, adding a new, previously unrecognized function of this accessory helicase.

scholarly journals Subcellular Localization and Rolling Circle Replication of Peach Latent Mosaic Viroid: Hallmarks of Group A Viroids

1999 ◽  
Vol 73 (8) ◽  
pp. 6353-6360 ◽  
Author(s):  
F. Bussière ◽  
J. Lehoux ◽  
D. A. Thompson ◽  
L. J. Skrzeczkowski ◽  
J.-P. Perreault
Keyword(s):  
Subcellular Localization ◽  
Cellular Level ◽  
Symmetric Mode ◽  
Rolling Circle Replication ◽  
Dot Blot ◽  
Rolling Circle ◽  
Group A ◽  
Group B ◽  
Contrast Group ◽  
Replication Intermediates

ABSTRACT We characterized the peach latent mosaic viroid (PLMVd) replication intermediates that accumulate in infected peach leaves and determined the tissue and subcellular localization of the RNA species. Using in situ hybridization, we showed that PLMVd strands of both plus and minus polarities concentrate in the cells forming the palisade parenchyma. At the cellular level, PLMVd was found to accumulate predominantly in chloroplasts. Northern blot analyses demonstrated that PLMVd replicates via a symmetric mode involving the accumulation of both circular and linear monomeric strands of both polarities. No multimeric conformer was detected, indicating that both strands self-cleave efficiently via their hammerhead sequences. Dot blot hybridizations revealed that PLMVd strands of both polarities accumulate equally but that the relative concentrations vary by more than 50-fold between peach cultivars. Taken together these results establish two hallmarks for the classification of viroids. Group A viroids (e.g., PLMVd), which possess hammerhead structures, replicate in the chloroplasts via the symmetric mode. By contrast, group B viroids, which share a conserved central region, replicate in the nucleus via an asymmetric mechanism. This is an important difference between self-cleaving and non-self-cleaving viroids, and the implications for the evolutionary origin and replication are discussed.

scholarly journals Autonomous Replication of the Conjugative Transposon Tn916

2016 ◽  
Vol 198 (24) ◽  
pp. 3355-3366 ◽  
Author(s):  
Laurel D. Wright ◽  
Alan D. Grossman
Keyword(s):  
Gene Transfer ◽  
Horizontal Gene Transfer ◽  
Origin Of Replication ◽  
Rolling Circle Replication ◽  
Conjugative Transposon ◽  
Rolling Circle ◽  
Autonomous Replication ◽  
Integrative And Conjugative Elements ◽  
Rolling Circle Mechanism ◽  
Origin Of Transfer

ABSTRACTIntegrative and conjugative elements (ICEs), also known as conjugative transposons, are self-transferable elements that are widely distributed among bacterial phyla and are important drivers of horizontal gene transfer. Many ICEs carry genes that confer antibiotic resistances to their host cells and are involved in the dissemination of these resistance genes. ICEs reside in host chromosomes but under certain conditions can excise to form a plasmid that is typically the substrate for transfer. A few ICEs are known to undergo autonomous replication following activation. However, it is not clear if autonomous replication is a general property of many ICEs. We found that Tn916, the first conjugative transposon identified, replicates autonomously via a rolling-circle mechanism. Replication of Tn916was dependent on the relaxase encoded byorf20of Tn916. The origin of transfer of Tn916,oriT(916), also functioned as an origin of replication. Using immunoprecipitation and mass spectrometry, we found that the relaxase (Orf20) and the two putative helicase processivity factors (Orf22 and Orf23) encoded by Tn916likely interact in a complex and that the Tn916relaxase contains a previously unidentified conserved helix-turn-helix domain in its N-terminal region that is required for relaxase function and replication. Lastly, we identified a functional single-strand origin of replication (sso) in Tn916that we predict primes second-strand synthesis during rolling-circle replication. Together these results add to the emerging data that show that several ICEs replicate via a conserved, rolling-circle mechanism.IMPORTANCEIntegrative and conjugative elements (ICEs) drive horizontal gene transfer and the spread of antibiotic resistances in bacteria. ICEs reside integrated in a host genome but can excise to create a plasmid that is the substrate for transfer to other cells. Here we show that Tn916, an ICE with broad host range, undergoes autonomous rolling-circle replication when in the plasmid form. We found that the origin of transfer functions as a double-stranded origin of replication and identified a single-stranded origin of replication. It was long thought that ICEs do not undergo autonomous replication. Our work adds to the evidence that ICEs replicate autonomously as part of their normal life cycle and indicates that diverse ICEs use the same replicative mechanism.

scholarly journals oriD structure controls RepD initiation during rolling-circle replication

2018 ◽  
Vol 8 (1) ◽  
Author(s):  
Algirdas Toleikis ◽  
Martin R. Webb ◽  
Justin E. Molloy
Keyword(s):  
Rolling Circle Replication ◽  
Rolling Circle

scholarly journals Rolling circle replication of DNA in yeast mitochondria.

1991 ◽  
Vol 10 (12) ◽  
pp. 3923-3929 ◽  
Author(s):  
R. Maleszka ◽  
P.J. Skelly ◽  
G.D. Clark-Walker
Keyword(s):  
Yeast Mitochondria ◽  
Rolling Circle Replication ◽  
Rolling Circle

Novel circular DNA virus identified in Opuntia discolor (Cactaceae) that codes for proteins with similarity to those of geminiviruses

2021 ◽  
Vol 102 (11) ◽  
Author(s):  
Rafaela S. Fontenele ◽  
Matias Köhler ◽  
Lucas C. Majure ◽  
Jesús A. Avalos-Calleros ◽  
Gerardo R. Argüello-Astorga ◽  
...  
Keyword(s):  
South America ◽  
Rolling Circle Replication ◽  
Dna Virus ◽  
Single Stranded Dna ◽  
Dna Viruses ◽  
Circular Dna ◽  
Rolling Circle ◽  
Rep Protein ◽  
Molecular Features ◽  
Viral Components

Viral metagenomic studies have enabled the discovery of many unknown viruses and revealed that viral communities are much more diverse and ubiquitous than previously thought. Some viruses have multiple genome components that are encapsidated either in separate virions (multipartite viruses) or in the same virion (segmented viruses). In this study, we identify what is possibly a novel bipartite plant-associated circular single-stranded DNA virus in a wild prickly pear cactus, Opuntia discolor, that is endemic to the Chaco ecoregion in South America. Two ~1.8 kb virus-like circular DNA components were recovered, one encoding a replication-associated protein (Rep) and the other a capsid protein (CP). Both of the inferred protein sequences of the Rep and CP are homologous to those encoded by members of the family Geminiviridae. These two putatively cognate components each have a nonanucleotide sequence within a likely hairpin structure that is homologous to the origins of rolling-circle replication (RCR), found in diverse circular single-stranded DNA viruses. In addition, the two components share similar putative replication-associated iterative sequences (iterons), which in circular single-stranded DNA viruses are important for Rep binding during the initiation of RCR. Such molecular features provide support for the possible bipartite nature of this virus, which we named utkilio virus (common name of the Opuntia discolor in South America) components A and B. In the infectivity assays conducted in Nicotiana benthamiana plants, only the A component of utkilio virus, which encodes the Rep protein, was found to move and replicate systemically in N. benthamiana. This was not true for component B, for which we did not detect replication, which may have been due to this being a defective molecule or because of the model plants (N. benthamiana) used for the infection assays. Future experiments need to be conducted with other plants, including O. discolor, to understand more about the biology of these viral components.

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