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SLIC-CAGE (Super-Low Input Carrier-CAGE) development: comparison with Saccharomyces cerevisiae nAnTi-CAGE and nanoCAGE libraries and validation of SLIC-CAGE on Mus musculus total RNA

ABSTRACT: SLIC-CAGE (Super-Low Input Carrier-CAGE) development: comparison with Saccharomyces cerevisiae nAnTi-CAGE and nanoCAGE libraries and validation of SLIC-CAGE on Mus musculus total RNA

PROVIDER: PRJEB25104 | ENA |

REPOSITORIES: ENA

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SLIC-CAGE (Super-Low Input Carrier-CAGE) development: comparison with Saccharomyces cerevisiae nAnTi-CAGE and nanoCAGE libraries and validation of SLIC-CAGE on Mus musculus total RNA

Project description:We developed SLIC-CAGE (Super-Low Input Carrier-CAGE) approach to capture 5'end of RNA polymerase II transcripts from as little as 5-10 ng of total RNA. The dramatic increase in sensitivity compared to existing CAGE methods is achieved by specially designed, selectively degradable carrier RNA. We tested SLIC-CAGE on Saccharomyces cerevisiae (BY4741 strain) and produced libraries from 1-100 ng of total cellular RNA. We also produced S. cerevisiae nAnT-iCAGE libraries as the current gold-standard CAGE libraries using the recommended 5 micrograms of total cellular RNA to assess the quality of SLIC-CAGE libraries produces with up to 1000-fold less material. We provide a direct comparison between SLIC-CAGE and the latest nanoCAGE protocol (libraries created using S. cerevisiae total RNA) and show that SLIC-CAGE produces unbiased libraries of higher complexity and quality than nanoCAGE. Finally, we provide SLIC-CAGE libraries on mouse embryonic stem cells (E14) using 5-100 ng of total cellular RNA as starting material.

2018-10-25 | E-MTAB-6519 | biostudies-arrayexpress

SLIC-CAGE (Super-Low Input Carrier-CAGE) application on mouse primordial germ cells isolated from embryonic day E11.5

2018-10-25 | E-MTAB-7056 | biostudies-arrayexpress

SLIC-CAGE (Super-Low Input Carrier-CAGE) application on mouse primordial germ cells isolated from embryonic day E11.5

Project description:SLIC-CAGE (Super-Low Input Carrier-CAGE) application on mouse primordial germ cells isolated from embryonic day E11.5

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SLIC-CAGE (Super-Low Input Carrier-CAGE) transcription start site mapping of mouse migratory and gonadal primordial germ cells, postnatal oocytes and early embryo

Project description:We used SLIC-CAGE to map transcription start sites (TSSs) of mouse primordial germ cells from embryonic days 9.5-16.5, postnatal oocytes (P6, P14 and MII), and early 2-cell and 4-cell mouse embryos. We use this TSS data to show that the mouse germline development starts with the somatic promoter code with a prominent switch to the maternal code (W-box dependent) occurring during the follicular oogenesis. We also find that the promoters of gonadal germ cells are characterised by a previously unknown divergence from the somatic transcription initiation. This divergence is distinct from the promoter code used later by the developing oocytes and reveals genome-wide promoter remodelling during early female and male germline development.

2022-02-14 | E-MTAB-9757 | biostudies-arrayexpress

Nuclear SLIC-CAGE of human cortical neurons from Cornelia de Lange Syndrome patients and control individuals

Project description:We used SLIC-CAGE to map transcriptional start sites in cortical neurons from Cornelia de Lange Syndrome (CdLS) patients and control individuals. SLIC-CAGE was performed using nuclear RNA isolated from pre-frontal cortical grey matter. Usage of nuclear RNA allows enrichment of unstable RNAs, such as RNA originating from enhancer transcription. We characterised promoter-level gene expression in cortical neurons from CdLS patients and found deregulation of hundreds of genes enriched for neuronal functions.

2021-02-27 | E-MTAB-9045 | biostudies-arrayexpress

SLIC-CAGE (Super-Low Input Carrier-CAGE) transcription start site mapping of mouse migratory and gonadal primordial germ cells, postnatal oocytes and early embryo

Project description:SLIC-CAGE (Super-Low Input Carrier-CAGE) transcription start site mapping of mouse migratory and gonadal primordial germ cells, postnatal oocytes and early embryo

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Nuclear SLIC-CAGE of human cortical neurons from Cornelia de Lange Syndrome patients and control individuals

Project description:Nuclear SLIC-CAGE of human cortical neurons from Cornelia de Lange Syndrome patients and control individuals

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Bio-carrier metagenome

Project description:Bio-carrier metagenome Metagenome

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Pig macrophage CAGE and RNA-seq libraries

Project description:Pig CAGE and macrophage-expressed genes in the pig genome

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Validation of picogram- and femtogram-input DNA libraries for microscale metagenomics.

Project description:High-throughput sequencing libraries are typically limited by the requirement for nanograms to micrograms of input DNA. This bottleneck impedes the microscale analysis of ecosystems and the exploration of low biomass samples. Current methods for amplifying environmental DNA to bypass this bottleneck introduce considerable bias into metagenomic profiles. Here we describe and validate a simple modification of the Illumina Nextera XT DNA library preparation kit which allows creation of shotgun libraries from sub-nanogram amounts of input DNA. Community composition was reproducible down to 100 fg of input DNA based on analysis of a mock community comprising 54 phylogenetically diverse Bacteria and Archaea. The main technical issues with the low input libraries were a greater potential for contamination, limited DNA complexity which has a direct effect on assembly and binning, and an associated higher percentage of read duplicates. We recommend a lower limit of 1 pg (?100-1,000 microbial cells) to ensure community composition fidelity, and the inclusion of negative controls to identify reagent-specific contaminants. Applying the approach to marine surface water, pronounced differences were observed between bacterial community profiles of microliter volume samples, which we attribute to biological variation. This result is consistent with expected microscale patchiness in marine communities. We thus envision that our benchmarked, slightly modified low input DNA protocol will be beneficial for microscale and low biomass metagenomics.

| S-EPMC5036114 | biostudies-literature

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