Ontology highlight
ABSTRACT:
PROVIDER: PRJNA699027 | ENA |
REPOSITORIES: ENA
Similar Datasets
Project description:We used the nanopore Cas9 targeted sequencing (nCATS) strategy to specifically sequence 125 L1HS-containing loci in parallel and measure their DNA methylation levels using nanopore long-read sequencing. Each targeted locus is sequenced at high coverage (~45X) with unambiguously mapped reads spanning the entire L1 element, as well as its flanking sequences over several kilobases. The genome-wide profile of L1 methylation was also assessed by bs-ATLAS-seq in the same cell lines (E-MTAB-10895).
2023-09-22 | E-MTAB-12247 | biostudies-arrayexpress
Project description:This dataset contains Xdrop followed by oxford nanopore long read sequencing performed in target tRNA gene deletion clones in HAP1 (t72) and HepG2 (t15). By applying de novo assembly based approach to Xdrop-LRS data, we identified Cas9-induced on-target genomic alteration.
2022-09-16 | E-MTAB-10652 | biostudies-arrayexpress
Project description:This dataset contains Xdrop followed by oxford nanopore long read sequencing performed in target tRNA gene deletion (t8) and intergenic region deletion (i50) clones in HepG2 . By applying de novo assembly based approach to Xdrop-LRS data, we identified Cas9-induced on-target genomic alteration.
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Project description:We used bs-ATLAS-seq to comprehensively map the genomic location and assess the DNA methylation status of human full-length LINE-1 elements (L1) in the genome of 2102Ep cells (E-MTAB-10895). We also achieved targeted nanopore sequencing to assay DNA methylation over a subset of loci (E-MTAB-12247). To further study the link between L1 DNA methylation and expression, we performed, in the same cell line, RNA-seq (E-MTAB-12246), as well as YY1 and H3K4me3 ChIP-seq (this dataset).
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Project description:Revertant mosaicism (RM) is a phenomenon in which inherited mutations are spontaneously corrected in somatic cells. RM occurs in some congenital skin diseases, although genetic validation of RM in clinically revertant skin has been challenging, especially when homologous recombination (HR) is responsible for RM. We introduced Nanopore Cas9-targeted sequencing (nCATS) for identifying HR in clinically revertant skin.
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Project description:The mammalian genome has an abundance of transposable elements, but understanding their contribution to complex biological systems is poorly understood. Here, we report the CRISPR/Cas9 deletion of a retrotransposon (Lx9c11) in mice and its effect on the immune response to virus infection. The regulatory role for Lx9c11 was assessed by RNA-seq analysis of knockout and wild-type mice, treated with CVB4 virus. Long read nanopore RNA-seq of transcripts enriched from Slfn1 locus (mm10, chr11:83109157-83116657).
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Project description:The CRISPR-Cas9 system enables efficient sequence-specific mutagenesis for creating germline mutants of model organisms. Key constraints in vivo remain the expression and delivery of active Cas9-guideRNA ribonucleoprotein complexes (RNPs) with minimal toxicity, variable mutagenesis efficiencies depending on targeting sequence, and high mutation mosaicism. Here, we established in vitro-assembled, fluorescent Cas9-sgRNA RNPs in stabilizing salt solution to achieve maximal mutagenesis efficiency in zebrafish embryos. Sequence analysis of targeted loci in individual embryos reveals highly efficient bi-allelic mutagenesis that reaches saturation at several tested gene loci. Such virtually complete mutagenesis reveals preliminary loss-of-function phenotypes for candidate genes in somatic mutant embryos for subsequent generation of stable germline mutants. We further show efficient targeting of functional non-coding elements in gene-regulatory regions using saturating mutagenesis towards uncovering functional control elements in transgenic reporters and endogenous genes. Our results suggest that in vitro assembled, fluorescent Cas9-sgRNA RNPs provide a rapid reverse-genetics tool for direct and scalable loss-of-function studies beyond zebrafish applications.
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2024-08-19 | E-MTAB-14368 | biostudies-arrayexpress
Project description:We sequenced DNA from the leaves of ten Col x Ler F1 hybrid plants (WT and recq4) using Nanopore long-read sequencing and identified crossover sites with COmapper. These data were used as a negative control for COmapper, as no crossover sites were expected to be detected. For nanopore sequencing of gDNA from leaves, leaves from 10 5-week-old plants were ground in liquid nitrogen using a mortar and pestle. The ground tissue was resuspended in four volumes of CTAB buffer (1% [w/v] CTAB, 50 mM Tris-HCl pH 8.0, 0.7 M NaCl, 10 mM EDTA) and incubated at 65°C for 30 min. Following chloroform extraction, isopropanol precipitation and removal of RNAs as above, the gDNA pellet was resuspended in 150 μl TE (10 mM Tris-HCl pH 8.0, 0.1 mM EDTA) buffer and gDNA was quantified using a Qubit dsDNA Broad Range assay kit (Thermo Fisher, Q32853). Nine micrograms of gDNA from pollen or seedlings was used to construct a nanopore long-read sequencing library using a Ligation Sequencing Kit V14 (Nanopore, SQK-LSK114). The libraries were sequenced using a PromethION platform (BGI, Hong Kong).
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