MobiChIP: a compatible library construction method of single-cell ChIP-seq based droplets
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ABSTRACT:
PROVIDER: PRJNA1141474 | ENA |
REPOSITORIES: ENA
Dataset's files
| Action | DRS | |||
|---|---|---|---|---|
| SRR30022371_1.fastq.gz | Fastqsanger.gz | |||
| SRR30022371_2.fastq.gz | Fastqsanger.gz | |||
| SRR30022372_1.fastq.gz | Fastqsanger.gz | |||
| SRR30022372_2.fastq.gz | Fastqsanger.gz | |||
| SRR30022373_1.fastq.gz | Fastqsanger.gz |
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Project description:In order to illustrate the epigenetic heterogeneity, versatile tools of single-cell ChIP-seq(scChIP-seq) are necessary to meet the convenience and accuracy. Here, we develop MobiChIP, a compatible ChIP-seq library construction method based current sequencing platform with single cell level. As a novel capture strategy, MobiChIP is efficient to capture the fragments from tagmented nuclei of numerous species and execute the mixing of samples from different tissues or species. Especially, this strategy enables the flexible sequencing manipulation and sufficient nucleosome amplification without customized sequencing primers. MobiChIP reveals the landscape of chromatin regulation regions with active(H3K27ac) and repressive(H3K27me3) histone modification markers in peripheral blood mononuclear cells (PBMCs), and accurately unveiled the epigenetic repression of hox gene cluster in PBMCs than ATAC-seq. Meanwhile, we complete the bioinformatics pipeline to integrate the scChIP-seq and scRNA-seq data to illustrate the cellular epigenetic and genetic heterogeneity.
2024-10-27 | GSE273350 | GEO
Project description:Next-generation sequencing has been widely used for the genome-wide profiling of histone modifications, transcription factor binding and gene expression through chromatin immunoprecipitated DNA sequencing (ChIP-seq) and cDNA sequencing (RNA-seq). Here, we describe a versatile library construction method that can be applied to both ChIP-seq and RNA-seq on the widely used Illumina platforms. Standard methods for ChIP-seq library construction require nanograms of starting DNA, substantially limiting its application to rare cell types or limited clinical samples. By minimizing the DNA purification steps that cause major sample loss, our method achieved a high sensitivity in ChIP-seq library preparation. Using this method, we achieved the following: (1) generated high-quality epigenomic and transcription factor-binding maps using ChIP-seq for murine adipocytes; (2) successfully prepared a ChIP-seq library from as little as 25 pg of starting DNA; (3) achieved paired-end sequencing of the ChIP-seq libraries; (4) systematically profiled gene expression dynamics during murine adipogenesis using RNA-seq; and (5) preserved the strand specificity of the transcripts in RNA-seq. Given its sensitivity and versatility in both double-stranded and single-stranded DNA library construction, this method has wide applications in genomic, epigenomic, transcriptomic and interactomic studies.
2015-12-15 | GSE75966 | GEO
Project description:Next-generation sequencing has been widely used for the genome-wide profiling of histone modifications, transcription factor binding and gene expression through chromatin immunoprecipitated DNA sequencing (ChIP-seq) and cDNA sequencing (RNA-seq). Here, we describe a versatile library construction method that can be applied to both ChIP-seq and RNA-seq on the widely used Illumina platforms. Standard methods for ChIP-seq library construction require nanograms of starting DNA, substantially limiting its application to rare cell types or limited clinical samples. By minimizing the DNA purification steps that cause major sample loss, our method achieved a high sensitivity in ChIP-seq library preparation. Using this method, we achieved the following: (1) generated high-quality epigenomic and transcription factor-binding maps using ChIP-seq for murine adipocytes; (2) successfully prepared a ChIP-seq library from as little as 25 pg of starting DNA; (3) achieved paired-end sequencing of the ChIP-seq libraries; (4) systematically profiled gene expression dynamics during murine adipogenesis using RNA-seq; and (5) preserved the strand specificity of the transcripts in RNA-seq. Given its sensitivity and versatility in both double-stranded and single-stranded DNA library construction, this method has wide applications in genomic, epigenomic, transcriptomic and interactomic studies. Pre-adipocytes and mature adipocytes were collected. Their chromatin and RNA were subjected to ChIP and mRNA extraction. Sequencing libraries from ChIP DNA or mRNA were generated following either standard protocols or TELP method. The quality and features of TELP libraries were proved and demonstrated in comparison with standard libraries or other published data.
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Project description:Background: Next-generation sequencing (NGS) is fundamental to the current biological and biomedical research. Construction of sequencing library is a key step of NGS. Therefore, various library construction methods have been explored. However, the current methods are still limited by some shortcomings. Results: This study developed a new NGS library construction method, Single strand Adaptor Library Preparation (SALP), by using a novel single strand adaptor (SSA). SSA is a double-stranded oligonucleotide with a 3ʹ overhang of 3 random nucleotides, which can be efficiently ligated to the 3′ end of single strand DNA by T4 DNA ligase. SALP can be started with any denatured DNA fragments such as those sheared by Tn5 tagmentation, enzyme digestion and sonication. When started with Tn5-tagmented chromatin, SALP can overcome a key limitation of ATAC-seq and become a high-throughput NGS library construction method, SALP-seq, which can be used to comparatively characterize the chromatin openness state of multiple cells unbiasly. In this way, this study successfully characterized the comparative chromatin openness states of four different cell lines, including GM12878, HepG2, HeLa and 293T, with SALP-seq. Similarly, this study also successfully characterized the chromatin openness states of HepG2 cells with SALP-seq by using 105 to 500 cells. Conclusions: This study developed a new NGS library construction method, SALP, by using a novel kind of single strand adaptor (SSA), which should has wide applications in the future due to its unique performance.
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Project description:In this study, we developed a novel plate-based 10X-compatible (PB10X) scRNA-seq strategy. By leveraging the Smart-seq3xpress (SS3X) principle, PB10X supports single-cell indexed sorting and cDNA generation that is downstream compatible with any commercially available 10X Single Cell 5' library construction kits such as the 10X Single Cell 5' Gene Expression and 5' V(D)J library construction kits. We demonstrated PB10X's performance on Jurkat T lymphoblasts by generating gene expression and V(D)J sequencing libraries, benchmarking it against 10X and SS3X.
2025-10-14 | GSE294062 | GEO
Project description:RNA modifications have important function in nearly every aspect of RNA biology. Epitranscriptome RNA modifications detected by high-throughput sequencing technologies is critical for understanding the distribution, regularity, stoichiometry, function of RNA modifications. One of disadvantages about these high-throughput sequencing technologies is needing high-input RNA. Here we use poly(A) tailing, template switching by reverse transcription during DNA synthesis and in vitro transcription (IVT) to develop a cutting-edge library construction strategy for profiling ultra-limited RNA modifications (Uli-Epic). We apply Uli-Epic for RNA-seq using 100 pg of poly A+ RNA. We also apply Uli-Epic to combine with BID-seq to characterize dynamics of pseudouridine in neural stem cells (NSC) and sperm under normal and fetal growth restriction (FGR) mice using as low as 100 pg of rRNA-depleted RNA. In addition, we combined Uli-Epic and GLORI to quantify the m6A methylomes of sperm and NSC under normal and FGR mice using 10ng of rRNA-depleted RNA. Summary, Uli-Epic is a strand-specific, high throughput and low input RNA library construction strategy which is compatible with many RNA modifications detected technologies.
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Project description:RNA modifications have important function in nearly every aspect of RNA biology. Epitranscriptome RNA modifications detected by high-throughput sequencing technologies is critical for understanding the distribution, regularity, stoichiometry, function of RNA modifications. One of disadvantages about these high-throughput sequencing technologies is needing high-input RNA. Here we use poly(A) tailing, template switching by reverse transcription during DNA synthesis and in vitro transcription (IVT) to develop a cutting-edge library construction strategy for profiling ultra-limited RNA modifications (Uli-Epic). We apply Uli-Epic for RNA-seq using 100 pg of poly A+ RNA. We also apply Uli-Epic to combine with BID-seq to characterize dynamics of pseudouridine in neural stem cells (NSC) and sperm under normal and fetal growth restriction (FGR) mice using as low as 100 pg of rRNA-depleted RNA. In addition, we combined Uli-Epic and GLORI to quantify the m6A methylomes of sperm and NSC under normal and FGR mice using 10ng of rRNA-depleted RNA. Summary, Uli-Epic is a strand-specific, high throughput and low input RNA library construction strategy which is compatible with many RNA modifications detected technologies.
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