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Sample GSM3511340 Query DataSets for GSM3511340
Status Public on Dec 14, 2018
Title wt_1_FeFREE
Sample type SRA
Source name germinated condia
Organism Neurospora crassa
Characteristics genotype: wt
growth protocol: iron-free condition
tissue: mycelia
Treatment protocol Iron-free cultures were grown in media without iron. All glassware including the one used to make the Vogel media was treated with 1 mM EDTA overnight, then with 5% HCl overnight, then rinsed well with Millipore water and baked
Growth protocol Cultures (2.5 x 10E6 conidia) were grown in standard Vogels medium and 1.5% sucrose (standard minimal medium) or in Iron-free Vogles and 1.5% sucrose, shaking for 48 hours at 32C.
Extracted molecule polyA RNA
Extraction protocol Mycelia were harvested and added to a tube containing ~300uL glass beads (260-300 micrometer, acid washed [Sigma Aldrich] and resuspended in sdH2O), 350uL NETS buffer (10mM Tris pH 7.5, 300mM NaCl, 1mM EDTA, 0.2% SDS) and 350uL phenol:chloroform:isoamyl alcohol (25:24:1) and lysed by a bead beater for 10 minutes at room temperature. Samples were cleared by centrifugation, and the aqueous (top) phase was added to 650uL cold 100% EtOH. Samples were precipitated at -20C, pelleted by centrifugation, washed twice with 1 ml 70% EtOH, and resuspended in DEPC-treated dH2O.
10 ug RNA was treated with DNAse (DNA-free™ DNA Removal Kit, Thermo Fisher Scientific), cleaned (Agencourt® RNAclean XP® beads, Beckman Coulter), and 4 ug DNAse-free RNA was used for RNA-seq libraries (KAPA Stranded mRNA-seq kit, KAPA Biosystems)
Library strategy RNA-Seq
Library source transcriptomic
Library selection cDNA
Instrument model Illumina HiSeq 4000
Data processing The sequencing data were uploaded to the Galaxy web platform, and the public server at was used to analyze the data. Specifically, the read quality was checked using FastQC, trimmed the adaptors using Trim Galore, aligned the reads and obtain read count using RNA STAR with intron length set to max 1000 bp. Differential gene expression analysis done by DESeq2 examined pair-wise differences between WT and Δve-1 after 48 hour growth under standard or iron-free conditions. Genes with log2 ≥ 1.0 or ≤ −1.0 changed expression and adjusted P values ≤ 0.05 were used for further analysis.
Genome_build: Neurospora crassa assembly 12 Fixed (files: neurospora_crassa_or74a_12_genome_FIXED.fasta, and neurospora_crassa_or74a_12_transcripts_FIXED.gtf; files are found in GEO submission GSE71024)
Supplementary_files_format_and_content: The .txt files were generated by the program RNA STAR at Galaxy platform. These RNASTAR.txt files report read number counts per gene for each replicate sample (two replicates total) from wild-type and ve-1 under standard or iron-free conditions. The four dataset.xlsx files were produced using the replicate RNASTAR.txt files in the program DESeq2 to determine genes that were differentially expressed > two fold and were significant (p < 0.05). Comparisons of mutant strains (N7373/N7374) versus WT (N3752/N3753) under standard (Dataset_1) and iron-free (Dataset_2) conditions as well as changes between standard and iron-free conditions in wt (Dataset_3) or ve-1 (Dataset-4) were created.
Submission date Dec 13, 2018
Last update date Dec 14, 2018
Contact name Tereza Ormsby
Organization name Charles University in Prague
Street address Hlavova 2030
City Prague 2
ZIP/Postal code 128 00
Country Czech Republic
Platform ID GPL23150
Series (1)
GSE123783 The Neurospora crassa VE-1/VE-2/LAE-1 velvet complex controls development, secondary metabolism and light-dependent carotenoid biosynthesis
BioSample SAMN10588781
SRA SRX5128284

Supplementary file Size Download File type/resource
GSM3511340_TGTTCCGT_3752-1_wt_FeFREE_48h_RNASTAR_reads.txt.gz 39.4 Kb (ftp)(http) TXT
SRA Run SelectorHelp
Raw data are available in SRA
Processed data provided as supplementary file
Processed data are available on Series record

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