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Sample GSM1534089

Query DataSets for GSM1534089

Status

Public on Apr 02, 2015

Title

RNA-seq in htz-1 mutants, replicate 2

Sample type

SRA

Source name

L3 larvae

Organism

Caenorhabditis elegans

Characteristics

tissue: whole larvae
strain: VC2490
Stage: L3
genotype/variation: htz-1 mutants

Treatment protocol

Total RNA was extracted in TriPure Reagent according to manufacturer's protocol, treated with baseline-zero DNAse (Epicentre, Madison, WI, USA), and purified using the Qiagen Rneasy kit.

Growth protocol

Worms grown in liquid culture were bleached and then hatched in M9 for 24-42 hrs. Hatched L1 larvae were inoculated in a proper volume of liquid culture. Next day when larvae reached the L3 stage they were cleaned by M9 washes, sorted on a COPAS BIOSORT (Union Biometrica) to isolate non-GFP homozygous mutants, and collected by freezing in liquid nitrogen.

Extracted molecule

total RNA

Extraction protocol

Total RNA was used to make mRNA libraries using the TruSeq RNA Sample Preparation Kit according to the manufacturer's instructions (Illumina, Inc., San Diego, CA, USA).

Library strategy

RNA-Seq

Library source

transcriptomic

Library selection

cDNA

Instrument model

Illumina HiSeq 2000

Description

Sample name: htz1-1012_RNAseq_MC002
processed data file: N2_lin-35_htz-1_processed_RNAseq.xlsx

Data processing

ChIP-seq and RNA-seq reads were aligned to the WS220 assembly of the C. elegans genome using BWA (Li and Durbin, 2010) with default settings.
ChIP-seq data were normalized using BEADS algorithm (Cheung et al, 2011).
ChIP-seq signals were exported to bigWig tracks at native resolution.
RNA-seq tag counts for each gene were extracted from BAM alignment files using HTSeq method working in ”union” mode. An exon model based on WS235 genes (taken from Ensembl Gene 74 December 2013) was lifted over to ce10/WS220.
RNA-seq tag counts were used to build the expression matrix. Differential gene expression between N2, lin-35 mutant, and htz-1 mutant was tested using DESeq2
Genome_build: ce10/WS220
Genome_build: http://ws220.wormbase.org/
Genome_build: http://hgdownload.soe.ucsc.edu/goldenPath/ce10/bigZips/
Supplementary_files_format_and_content: "linear" scores in bigWig tracks represent BEADS values; files were generated using BEADS normalization pipeline in R
Supplementary_files_format_and_content: "zscore" scores in bigWig tracks represent standardized BEADS values; files were generated using BEADS normalization pipeline in R
Supplementary_files_format_and_content: xslx spreadsheet contains: reads per kilobase (kb) of transcript per millions reads in library (RPKM) for all RNA-seq experiments, averaged RPKM values for replicates, plus log2 fold changes, p-values and adjusted p-values (FDR) calculated with DEseq2 for gene knock-out replicates versus N2 (wild-type) replicate

Submission date

Oct 29, 2014

Last update date

May 15, 2019

Contact name

Przemyslaw Aleksander Stempor

Organization name

University of Cambridge

Department

The Gurdon Institute

Lab

Ahringer Lab