The Transposon Insertion site Profiling Chip (TIP-chip)
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ABSTRACT:
ORGANISM(S): Saccharomyces cerevisiae
PROVIDER: GSE5646 | GEO | 2007/01/01
SECONDARY ACCESSION(S): PRJNA96845
REPOSITORIES: GEO
Similar Datasets
Project description:We have developed a microarray intended for use in finding all transposons in a region of interest. By selectively amplifying and hybridizing transposon flanking DNA to our array, we can localize all transposons in the region present on our TIP-chip, a dense tiling array. We have tested our technology in yeast and have been successful. The two FY2 hybridizations are technical replicates and serve as a benchmark -- we compared these results to the published S288C sequence. The two GRF167 samples are also technical replicates and these are the base strain for the L27-10 hybridizations, and therefore serve as controls for those hybridizations. The three L27-10 hybridizations are all technical replicates.
2010-07-01 | E-GEOD-5646 | biostudies-arrayexpress
Project description:We characterized changes of transposon and mRNA expressions in armi, rhino ,aub, ago3 mutants with respect to wild type using Affy tiling array. In most of these mutants, mRNA expressions were mostly unchanged but increased expressions was observed for many transposons indicating the role of these proteins in silencing transposons in Drosophila ovaries Keywords: Tiling array transcriptome profiling
2009-05-03 | GSE14370 | GEO
Project description:Transposable genetic elements are ubiquitous, yet their presence or absence at any given position within a genome can vary between individual cells, tissues, or strains. Transposable elements have profound impacts on host genomes by altering gene expression, assisting in genomic rearrangements, causing insertional mutations, and serving as sources of phenotypic variation. Characterizing a genome?s full complement of transposons requires whole genome sequencing, precluding simple studies of the impact of transposition on interindividual variation. Here, we describe a global mapping approach for identifying transposon locations in any genome, using a combination of transposon-specific DNA extraction and microarray- based comparative hybridization analysis. We use this approach to map the repertoire of endogenous transposons in different laboratory strains of Saccharomyces cerevisiae and demonstrate that transposons are a source of extensive genomic variation. We also apply this method to mapping bacterial transposon insertion sites in a yeast genomic library. This unique whole genome view of transposon location will facilitate our exploration of transposon dynamics, as well as defining bases for individual differences and adaptive potential. Keywords: transposon mapping
2006-11-15 | GSE6278 | GEO
Project description:Lipid intermediates derived from sphingolipid metabolism are crucial regulators of mitochondrial function from yeast to humans. Among these intermediates, trans-2-hexadecenal (t-2hex) within the sphingolipid degradation pathway exhibits remarkable efficiency in inducing mitochondria-mediated cell death. In yeast cell cultures, the addition of t-2-hex triggers complete disintegration of the mitochondrial network, leading to subsequent cell death. This effect is particularly pronounced in yeast cells lacking the activity of the t-2-hex degrading enzyme, Hfd1. However, the molecular mechanisms of t-2-hex induction of mitochondrial dysfunction are completely unknown. In this project, we want to exploit the unprecedented power of yeast genetics to unveil novel genetic determinants involved in t-2-hex's pro-apoptotic function. To accomplish this, we employed the SATAY method, which combines saturated transposon mutagenesis with high-throughput sequencing to functionally explore the yeast genome. In our screening approach, hfd1 mutant cells harboring a plasmid-encoded inducible MiniDs transposon were induced by galactose, resulting in extensive integration of the transposon throughout the yeast genome. Cells with the plasmid excised and the transposon genomically integrated were pooled together, creating a high-density transposon library comprising approximately 2.3E+06 independent insertion mutants. Subsequently, the pooled mutant library was subjected to treatment with the mitochondria-mediated death inducer, t-2-hexadecenal. As a control, cells were also incubated with the solvent dimethyl sulfoxide (DMSO), in which hexadecenal is dissolved. Following the treatments, cells were collected for genomic DNA extraction and digestion, using restriction enzymes with frequent four-base pair recognition sites. The resulting library fragments were circularized using T4 DNA ligase, and the transposon-genome junctions were selectively amplified through PCR with outward-facing primers specific to the transposon. Finally, the pooled and purified amplicons were subjected to massive sequencing on an Illumina MiSeq platform. The obtained sequences were then aligned to the reference genome of Saccharomyces cerevisiae, allowing for the mapping of transposon insertions and the calculation of transposon counts per gene. This project enabled the identification of genes required for the resistance and toxicity to t-2hex.
2023-08-31 | E-MTAB-13208 | biostudies-arrayexpress
Project description:Transposon screens are powerful in vivo assays used to identify loci driving carcinogenesis. These loci are identified as Common Insertion Sites (CIS), i.e. regions with more transposon insertions than expected by chance. However, the identification of CIS is strongly affected by biases in the insertion behaviour of transposon systems. Here, we introduce Transmicron, a novel method that differs from previous methods by i) modelling neutral insertion rates based on chromatin accessibility, transcriptional activity, and sequence context, and ii) estimating oncogenic selection for each genomic region using Poisson regression to model insertion counts while controlling for neutral insertion rates. To assess the benefits of our approach, we generated a dataset applying two different transposon systems under comparable conditions. Benchmarking for enrichment of known cancer genes showed improved performance of Transmicron against state-of-the-art methods. Modelling neutral insertion rates allowed for better control of false positives and stronger agreement of the results between transposon systems. Moreover, using Poisson regression to consider intra-sample and inter-sample information proved beneficial in small and moderately-sized datasets. Transmicron is open-source and freely available. Overall, this study contributes to the understanding of transposon biology and introduces a novel approach to use this knowledge for discovering cancer driver genes.
2022-10-01 | GSE214379 | GEO
Project description:Streptococcus pneumoniae is a major cause of serious infections such as pneumonia and meningitis in both children and adults worldwide. Here, we describe the development of a high-throughput genome-wide technique, Genomic Array Footprinting (GAF), for the identification of genes essential for this bacterium at various stages during infection. GAF enables negative screens by means of a combination of transposon mutagenesis and microarray technology for the detection of transposon insertion sites. We tested several methods for the identification of transposon insertion sites and found that amplification of DNA adjacent to the insertion site by PCR resulted in non-reproducible results, even when combined with an adapter. However, restriction of genomic DNA followed directly by in vitro transcription circumvented these problems. Analysis of parallel reactions generated with this method on a large mariner transposon library, showed that it was highly reproducible and correctly identified essential genes. Comparison of a mariner library to one generated with the in vivo transposition plasmid pGh:ISS1, showed that both have an equal degree of saturation, but that 9% of the genome is preferentially mutated by either one. The usefulness of GAF was demonstrated in a screen for genes essential for survival of zinc stress. This identified a gene encoding a putative cation efflux transporter, and its deletion resulted in an inability to grow under high zinc conditions. In conclusion, we developed a fast, versatile, specific, and high-throughput method for the identification of conditionally essential genes in S. pneumoniae. Keywords: GAF Identification of transposon insertion sites
2007-06-26 | GSE6913 | GEO
Project description:The piRNA pathway controls transposon expression in animal germ cells, thereby ensuring genome stability over generations. piRNAs are maternally deposited and required for proper transposon silencing in adult offspring. However, a long-standing question in the field is the precise function of maternally deposited piRNAs and its associated factors during embryogenesis. Here, we probe the spatio-temporal expression patterns of several piRNA pathway components during early stages of development. Amongst those, factors required for transcriptional gene silencing (TGS) showed ubiquitous abundance in somatic and pole cells throughout the first half of embryogenesis. We further analysed the transcriptomes of various embryo stages and correlated these with the presence of selected chromatin marks. We found that a number of transposon families show bursts of transcription during early embryonic stages. Transposons heavily targeted by maternally deposited piRNAs accumulated repressive chromatin marks following their spike in expression. Furthermore, depletion of maternally deposited Piwi protein in early embryos resulted in increased expression of transposons targeted by inherited piRNAs and was accompanied by a strong loss of repressive chromatin marks at coding sequences. Overall, our data suggests a pivotal role for the piRNA pathway in transposon defence during Drosophila embryogenesis in somatic cells.
2021-07-26 | PXD022409 | Pride
Project description:Mobile elements are important components of our genomes, with diverse and significant effects on phenotype. Not only can transposons inactivate genes by direct disruption and shuffle the genome through recombination, they can also alter gene expression subtly or powerfully. Currently active transposons are highly polymorphic in host populations, including, among hundreds of others, L1 and Alu elements in humans and Ty1 elements in yeast. For this reason, we wished to develop a simple genome-wide method for identifying all transposons in any given sample. We have designed a transposon insertion site profiling chip (TIP-chip), a microarray intended for use as a high-throughput technique for mapping transposon insertions. By selectively amplifying transposon flanking regions and hybridizing them to the array, we can locate all transposons present in a sample. We have tested the TIP-chip extensively to map Ty1 retrotransposon insertions in yeast and have achieved excellent results in two laboratory strains as well as in evolved Ty1 high-copy strains. We are able to identify all of the theoretically detectable transposons in the FY2 lab strain, with essentially no false positives. In addition, we mapped many new transposon copies in the high-copy Ty1 strain and determined its Ty1 insertion pattern.
| S-EPMC1693798 | biostudies-literature
Project description:Bombyx Vasa (BmVasa) assembles non-membranous organelle, nuage or Vasa bodies, in germ cells, known as the center for Siwi-dependent transposon silencing and concomitant Ago3-piRISC biogenesis. However, details of the body assembly remain unclear. Here, we show that the N-terminal intrinsically disordered region (N-IDR) and RNA helicase domain of BmVasa are responsible for self-association and RNA binding, respectively, but N-IDR is also required for full RNA-binding activity. Both domains are essential for Vasa body assembly in vivo and droplet formation in vitro via phase separation. FAST-iCLIP reveals that BmVasa preferentially binds transposon mRNAs. Loss of Siwi function derepresses transposons but has marginal effects on BmVasa-RNA binding. This study shows that BmVasa assembles nuage by phase separation via its ability to self-associate and bind newly exported transposon mRNAs. This unique property of BmVasa allows transposon mRNAs to be sequestered and enriched in nuage, resulting in effective Siwi-dependent transposon repression and Ago3-piRISC biogenesis.
2023-03-06 | GSE213915 | GEO
Project description:Bombyx Vasa (BmVasa) assembles non-membranous organelle, nuage or Vasa bodies, in germ cells, known as the center for Siwi-dependent transposon silencing and concomitant Ago3-piRISC biogenesis. However, details of the body assembly remain unclear. Here, we show that the N-terminal intrinsically disordered region (N-IDR) and RNA helicase domain of BmVasa are responsible for self-association and RNA binding, respectively, but N-IDR is also required for full RNA-binding activity. Both domains are essential for Vasa body assembly in vivo and droplet formation in vitro via phase separation. FAST-iCLIP reveals that BmVasa preferentially binds transposon mRNAs. Loss of Siwi function derepresses transposons but has marginal effects on BmVasa-RNA binding. This study shows that BmVasa assembles nuage by phase separation via its ability to self-associate and bind newly exported transposon mRNAs. This unique property of BmVasa allows transposon mRNAs to be sequestered and enriched in nuage, resulting in effective Siwi-dependent transposon repression and Ago3-piRISC biogenesis.
2023-03-06 | GSE213914 | GEO