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Sample GSM6235097

Query DataSets for GSM6235097

Status

Public on Jun 16, 2022

Title

SN_0_7

Sample type

SRA

Source name

cultures

Organism

Mycosarcoma maydis

Characteristics

tissue: cultures
genotype: wild type R521
treatment: grown in supernatant of U.maydis cells treated with 0.7% H2O2

Growth protocol

Ustilago maydis wild type was grown for 16h in YEPS (1% (w/v) yeast extract, 2% (w/v) peptone, 2% (w/v) saccharose). The same amount of 2x107 cells of U.maydis wild type overnight culture was transfered to 10% YEPS (as control) as well as to cell-free supernatants of U.maydis wild type treated with H202 in two different concentrations (0.4% and 0.7%) (as two separate conditions).

Extracted molecule

total RNA

Extraction protocol

Total RNA was extracted using GeneJET RNA Purification kit (ThermoScientific, Lithuania) according to the manufacturer recommendation for yeast total RNA isolation. DNA was removed from total RNA sampels using DNAfree DNase treatment and Removal Kit (Ambion). Purified RNA was analzyed by RNA 6000 Nano assay on Agilent 2100 Bioanalyzer (Agilent Technologies, CA, USA). Samples were further processed by Novogene as follows: messenger RNA (mRNA) was purified from total RNA using poly-T oligo-attached magnetic beads.
After fragmentation, the first strand cDNA was synthesized using random hexamer primers, followed by the second strand cDNA synthesis using dTTP. The library was checked with Qubit and real-time PCR for quantification and bioanalyzer for size distribution detection. Quantified libraries were pooled and sequenced on Illumina NovaSeq 6000 platform using 150bp paired-end sequencing.

Library strategy

RNA-Seq

Library source

transcriptomic

Library selection

cDNA

Instrument model

Illumina NovaSeq 6000

Data processing

Raw FASTQ data were preprocessed using fastp to remove adapters, poly-N>10% sequences and low quality reads (Qscore of over 50% bases of the read <=5). Clean data served for calculating Q20, Q30 and GC content.
Paired-end clean reads were mapped to the reference genome using HISAT2 software.
Featurecounts was used to count reads mapped for each gene and expressed in FPKM (fragments per kilobase of exone model per million mapped reads). Prior to differential gene expression analysis, for each sequenced library, the read counts were adjusted by Trimmed Mean- of M-values (TMM) through one scaling normalized factor. Differential expression analysis of two conditions was performed using EdgeR (without biological replicates) and DESeq2 (with three biological replicates per condition) R packages. The p-values were adjusted using Benjamini and Hochberg approach for controlling the false discovery rate. Genes with adjusted p-value < 0.005 and |log2(fold Change)| >1 found by EdgeR were assigned as differential expressed.
Assembly: RefSeq GCF_000328475.2 Genome assembly Umaydis521_2.0 (https://www.ncbi.nlm.nih.gov/data-hub/genome/GCF_000328475.2/)
Supplementary files format and content: Excel files including FPKM values for each sample in three comparisons: 1) Differential gene expression analysis between U.maydis control and U.maydis grown in supernatant of U.maydis cells treated with 0.4% H2O2 (SN_0_4vsControl_DEG_all), 2)Differential gene expression analysis between U.maydis control and U.maydis grown in supernatant of U.maydis cells treated with 0.7% H2O2 (SN_0_7vsControl_DEG_all) and 3) Differential gene expression analysis between U.maydis grown in supernatant of U. maydis cells treated with 0.7% H2O2 and in supernatant of U. maydis cells treated with 0.4% H2O2 (SN_0_7vsSN_0_4_DEG_all)

Submission date

Jun 11, 2022

Last update date

Jun 16, 2022

Contact name

Stefan Stanovčić

E-mail(s)

[email protected]

Phone

+381649485967

Organization name

IMGGE