Browse
Submit Data
Databases
API
Help

Dataset Information

0 Views

0 Connections

0 Citations

0 Reanalyses

0 Downloads

Omics score: 0

Sequencing of reaction products from protospacer integration by Cas1-Cas2

ABSTRACT: We report the sites of protospacer integration into plasmids by Cas1-Cas2. The goal of this study is to determine the role of sequence elements on protospacer integration.

ORGANISM(S): Cloning vector pUC19

PROVIDER: GSE64552 | GEO | 2015/01/30

SECONDARY ACCESSION(S): PRJNA271290

REPOSITORIES: GEO

ACCESS DATA

Json Xml

Dataset's files

Source:

			Action	DRS
		Other

Items per page:

1 - 1 of 1

Similar Datasets

Sequencing of reaction products from protospacer integration by Cas1-Cas2

Project description:We report the sites of protospacer integration into plasmids by Cas1-Cas2. The goal of this study is to determine the role of sequence elements on protospacer integration. Methods: Cas1-Cas2 integration products were fragmented, end-repaired, adapter ligated, PCR amplified, and analyzed by Illumina sequencing. Protospacer-containing reads were selected by Cutadapt and mapped to the plasmid with Bowtie.

2015-01-30 | E-GEOD-64552 | biostudies-arrayexpress

Protospacer integration into plasmid DNA by Cas1–Cas2

Project description:We report the integration site specificity of Cas1–Cas2 into target plasmid DNA encoding a CRISPR locus. The goal of this study is to determine the effect of IHF on target site selection of Cas1–Cas2.

2016-05-27 | GSE79992 | GEO

Cloning vector pUC19

Project description:Protospacer integration into plasmid DNA by Cas1–Cas2

| PRJNA317579 | ENA

Cloning vector pUC19

Project description:Sequencing of reaction products from protospacer integration by Cas1-Cas2

| PRJNA271290 | ENA

Transcription Termination and Antitermination of Bacterial CRISPR Arrays

Project description:V. cholerae A50 has a functional CRISPR-cas system with a conserved boxA sequence. Plasmids harboring protospacers that are perfect targets for each spacer of the array are introduced into wt and boxA mutant V. cholerae. After a period of growth without selection, cells are collected and the protospacer plasmids are sequenced in a high throughput manner.

2020-10-08 | E-MTAB-9631 | biostudies-arrayexpress

Vibrio Cholerae CRISPR-cas PAM requirements

Project description:Plasmids were constructed harboring protospacers that are perfect targets for spacers #4 and #21 of the array, but contain NNN (all possible PAM combinations) immediately upstream of the protospacer. The plasmids were introduced into V. cholerae with or without a functional CRISPR-cas system and cells were plated on selective media. Cells were collected and the protospacer plasmids were sequenced in a high throughput manner. PAMs were counted using a custom python script

2024-05-06 | E-MTAB-14044 | biostudies-arrayexpress

Cas1-Cas2 and DnaK physically and functionally interact to modulate CRISPR Adaptation

Project description:Prokaryotic Cas1-Cas2 protein complexes generate adaptive immunity to mobile genetic elements (MGEs), by capture and integration of MGE DNA in to CRISPR sites. De novo immunity relies on Cas1-Cas2 targeting MGE DNA, without the aid of pre-existing immunity complexes, through mechanisms of ‘naive adaptation’ that are not clear. Using E. coli we show that the chaperone DnaK inhibits Cas1-Cas2 from DNA binding and integration, and that DnaK expression prevents naïve adaptation from chromosomal self-targeting. We show that that inhibition of naïve adaptation is reversible by eliminating DnaK from cells, by mutation of the DnaK substrate binding domain, and by expression of an MGE (phage )protein. We show that a fluorescently labelled Cas1 fusion can be visualised in living cells. Formation of foci depends on active DNA replication, and that the number of foci per cell is much increased in cells lacking DnaK. We discuss a model in which DnaK provides a mechanism for restraining naïve adaptation from self-targeting. This restraint is released once MGE DNA is present in the cell.

2023-05-18 | PXD042090 | Pride

Casposase structure and the mechanistic link between DNA transposition and spacer acquisition by CRISPR-Cas

Project description:Key to CRISPR-Cas adaptive immunity is maintaining an ongoing record of invading nucleic acids that are encountered, a process carried out by the Cas1-Cas2 complex that integrates short segments of foreign genetic material (spacers) into the CRISPR locus. It is hypothesized that Cas1 evolved from casposases, a novel class of transposases. We show here that casposase integration in vitro recapitulates several properties of CRISPR-Cas integrases. The X-ray structure of Methanosarcina mazei casposase bound to DNA representing the product of integration reveals a tetramer with target DNA bound snugly between two dimers in which single-stranded casposon end binding resembles that of spacer 3'-overhangs. The differences between transposase and CRISPR-Cas integrase are largely architectural, and it appears that evolutionary change involved changes in protein-protein interactions to favor Cas2 binding over tetramerization and the separation of Cas1 dimers. This, in turn, led to preferred integration of single spacers over two transposon ends.

2020-02-03 | GSE139037 | GEO

Cas1 and Fen1 Display Commutable Functions During Archaeal DNA Damage Repair

Project description:CRISPR-Cas constitutes an adaptive prokaryotic defence system against invasive nucleic acids like viruses and plasmids. Beyond their role in immunity, CRISPR-Cas systems have been shown to closely interact with components of cellular DNA repair pathways either by regulating their expression or via direct protein-protein contact or enzymatic activity. The integrase Cas1 is usually involved in the adaptation phase of CRISPR-Cas immunity but its function in cellular DNA repair pathways has been proposed before. Here, we analysed the capacity of an archaeal Cas1 from Haloferax volcanii to compensate DNA damage induced by oxidative stress and found that a deletion of the cas1 gene led to severe growth defects after stress induction. In addition, our results indicate that Cas1 is directly involved in DNA damage repair as the enzymatically active site of the protein is crucial for growth rescue under oxidative conditions. Based on biochemical cleavage assays, we propose a mechanism in which Cas1 exerts a similar function like the DNA repair protein Fen1 by resolving branched repair intermediate structures. Overall, the present study broadens our understanding of the functional link between CRISPR-Cas immunity and DNA repair by demonstrating that Cas1 and Fen1 display commutable roles during archaeal DNA damage repair.

2022-05-20 | PXD029952 | Pride

Sequence Requirements for Cascade Function in a Type I-E CRISPR System: Plasmid Loss

Project description:We generated a collection of 13 plasmids, with each plasmid containing a variant of a CRISPR protospacer targeted by spacer 8 of the E. coli CRISPR-I array. We transformed the plasmids as a pool into delta cas3 E. coli cells expressing all other cas genes constitutively. We then transformed these cells with either an empty vector or a plasmid expressing the Cas3 nuclease. DNA surrounding the protospacers was PCR-amplified and sequenced.

2017-08-24 | E-MTAB-5969 | biostudies-arrayexpress

OmicsDI is part of the ELIXIR infrastructure

OmicsDI is an Elixir interoperability service. Learn more ›

Tweets

OmicsDI Databases

PRIDE
PeptideAtlas
MassIVE
JPOST Repository
Physiome Model Repository

EGA
EVA
ENA
LINCS
PAXDB
Cell Collective

MetaboLights
Metabolomics Workbench
MetabolomeExpress
GNPS
BioModels
FAIRDOMHub

ArrayExpress
dbGaP
ExpressionAtlas
GEO
NODE

Information

Databases
Help
API
Contact us
Code on GitHub
Terms of use
Submit Data