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To investigate 4-TU labelling kinetics in a novel zebrafish transgenic line (lf:UPRT). [Adult SLAM-Itseq]

ABSTRACT: To investigate 4-TU labelling kinetics in a novel zebrafish transgenic line (lf:UPRT). [Adult SLAM-Itseq]

PROVIDER: PRJNA962798 | ENA |

REPOSITORIES: ENA

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			Action	DRS
	SRR24331581.fastq.gz	Fastqsanger.gz
	SRR24331582.fastq.gz	Fastqsanger.gz
	SRR24331583.fastq.gz	Fastqsanger.gz
	SRR24331584.fastq.gz	Fastqsanger.gz
	SRR24331585.fastq.gz	Fastqsanger.gz

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To investigate 4-TU labelling kinetics in a novel zebrafish transgenic line (lf:UPRT). [Adult SLAM-Itseq]

Project description:To understand 4-TU labelling kinetics in a novel zebrafish transgenic line (lf:UPRT), we exposed adult lf:UPRT zebrafish (whereby animals have hepatocyte-specific uracil phosphoribosyltransferase expression) with 4-TU for 1, 3, 6 and 9 hours. We then dissected adult livers for SLAM-ITseq to determine the optimal 4-TU labelling time in our SLAM-ITseq workflow to allow us to study hepatocyte-specific nascent transcriptional changes.

2024-01-13 | GSE230830 | GEO

SLAM-ITseq identifies that Nrf2 induces liver regeneration through the pentose phosphate pathway [Larval SLAM-Itseq]

Project description:We investigated the immediate transcriptional changes in hepatocytes following APAP-induced acute liver injury by exposing lf:UPRT zebrafish larvae to DMSO or APAP in the presence of 4TU. We the performed gene expression profiling analysis on total and nascent counts using data obtained from SLAM-ITseq of WT and Nrf2KO animals.

2024-01-13 | GSE230831 | GEO

C. elegans L4 hypodermal SLAM-ITseq

Project description:To adapt SLAM-ITseq for Caenorhabditis elegans, we inserted a codon-optimized version of the Toxoplasma gondii UPRT gene downstream of a FRT-flanked mCh::his-58 cassette and under control of the strong and ubiquitous eft-3 promoter. We introduced a single copy of this construct into the C. elegans genome and crossed the resulting line with a dpy-7p::FLP driver. FLP-dependent expression of UPRT was confirmed by Western blotting (strain BN1190). As a proof of concept, we exposed animals to 4-TU from L3 to L4 stage and purified total RNA. RNA from animals without FLP expression was used to control for UPRT-independent labeling strain BN1158). RNA was alkylated with iodoacetamide, which leads to a T>C conversion of 4-TU-labeled positions during cDNA synthesis, and subjected to deep sequencing (see Materials and Methods). We confirmed that the presence of FLP and UPRT did not affect gene expression. Comparing the T>C conversion rate per gene revealed a significant increase in nematodes expressing UPRT in the hypodermis. Next, we performed a beta-binominal test to identify genes that were significantly labelled across all replicas, which retrieved 197 genes (p<0.003; FDR<0.1 after adjustment for multiple comparisons). Original SLAM-ITseq reference: Matsushima W, Herzog VA, Neumann T, Gapp K, Zuber J, Ameres SL, Miska EA (2018) SLAM-ITseq: sequencing cell type-specific transcriptomes without cell sorting. Development 145 (13). doi:10.1242/dev.164640

2022-12-31 | GSE192419 | GEO

SLAM-Seq analysis of Ythdf2-deficient pre-leukemic cells

Project description:The mRNA m6A reader YTHDF2 is overexpressed in a broad spectrum of human acute myeloid leukemias (AML). To study the role of YTHDF2 on mRNA decay rates in leukemia, c-Kit+ cells from foetal livers of Ythdf2fl/fl; Vav-iCre (Ythdf2CKO) and Ythdf2fl/fl (Ythdf2CTL) 14.5 dpc embryos were transduced with Meis1 and Hoxa9 oncogenes and serially re-plated to generate pre-leukemic cells. Medium with 4SU was used for pre-leukemic cells labelling for 12 hours and was later replaced with 4SU-free medium (time 0). Cells were collected immediately after medium change and at 1, 3 and 9 hours for library generation. RNA from Ythdf2CKO (n=3 biological replicates) and Ythdf2CTL (n=3 biological replicates) pre-leukemic cells were used for SLAM-seq library generation.

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Soil metagenome LF

Project description:Soil metagenome LF

| PRJNA931815 | ENA

Streptomyces violaceusniger Tu 4113 strain:Tu 4133

Project description:Streptomyces violaceusniger Tu 4113 genome sequencing project

| PRJNA43031 | ENA

Streptomyces sp. Tu 2975

Project description:Streptomyces sp. Tu 2975 Genome sequencing

| PRJNA590865 | ENA

Streptomyces collinus Tu 365

Project description:Streptomyces collinus Tu 365 Genome sequencing

| PRJNA171216 | ENA

TU-tagging: cell type specific RNA isolation from intact complex tissues

Project description:We show that the combination of spatially restricted uracil phosphoribosyltransferase (UPRT) expression with 4-thiouracil (4TU) delivery can be used to label and purify cell type specific RNA from intact complex tissues. This method is useful for isolating RNA from cell types that are difficult to isolate by dissection or dissociation methods and should work in any organism where UPRT can be spatially expressed, including mammals and other vertebrates. To confirm that thio-labeled RNA was from UPRT-expressing cells and demonstrate the utility of TU-tagging for cell type-specific RNA isolation, we purified TU-tagged and untagged RNA and compared them using microarrays. We chose to isolate RNA from larval glia, which are present in low number, are highly dispersed, and have a complex cell morphology, making them one of the most difficult cell types to isolate by dissection or dissociation methods. We used reversed polarity (repo)-GAL4 to drive expression of UAS-UPRT specifically in glial cells of the larval brain. We purified TU-tagged and untagged RNA from 72-96h larval brains, and hybridized them to custom Agilent microarrays. We performed two biological replicate experiments.

2010-06-19 | E-GEOD-15445 | biostudies-arrayexpress

Danio rerio breed:TU

Project description:Danio rerio breed:TU Raw sequence reads

| PRJNA599540 | ENA