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Sample GSM7334253

Query DataSets for GSM7334253

Status

Public on Oct 23, 2023

Title

KN99a_time_24h_replicate_2

Sample type

SRA

Source name

cell Culture

Organism

Cryptococcus neoformans

Characteristics

tissue: cell Culture
strain: KN99a
cell type: Yeast
time point: 24h

Treatment protocol

RNA was isolated from strains before or after growth for 24 hours in RPMI + 10% mouse serum (37°C, 5% CO2)

Growth protocol

Strains were streaked from storage at -80 °C onto YPD plates, incubated for two days at 30 °C, inoculated from single colonies into YPD liquid cultures, and grown at 30 °C with shaking (220 rpm)

Extracted molecule

total RNA

Extraction protocol

Approximately 2 × 10^8cells were collected by centrifugation, suspended in TRIzol reagent (from Invitrogen), and subjected to mechanical lysis by bead beating at 4°C with 0.5-mm glass beads for 1 min, followed by a 2-min rest, for a total of 5 cycles. Following lysis, total RNA was extracted according to the manufacturer's instructions. Residual DNA was removed from the RNA preparation by treatment with the Turbo DNA-free kit (from Ambion) according to the manufacturer's instructions.
We used the NEBNext Ultra Directional RNA Library Prep Kit for Illumina protocol according to the manufacturer's instruction.
NEBNext Ultra Directional RNA Library Prep Kit for Illumina

Library strategy

RNA-Seq

Library source

transcriptomic

Library selection

cDNA

Instrument model

Illumina NextSeq 500

Data processing

Read quality was measured with FastQC
Fastq files were aligned to the KN99a genome ASM14924v2 using Hisat2 version 2.2.1 with the default parameters plus --max-intronlen 2200
SAM files were converted to bam, reads were sorted and indexed, and read duplicates were removed from the final bam files using SAMtools 1.7
d and indexed, and read duplicates were removed from the final bam files using SAMtools 1.7.
The number of reads mapped per gene was calculated using featureCounts from the package Subread 2.0.0
Assembly: Differential gene expression was analyzed with DESeq2, using the IHW (independent hypothesis weighting) package to calculate adjusted p-values
Supplementary files format and content: C8.24_KN99.24.csv
Supplementary files format and content: Table containing comparison of gene expression between C8 and KN99a (C8/KN99a) after growth for 24 hours in RPMI + 10% mouse serum (37°C, 5% CO2)

Submission date

May 12, 2023

Last update date

Oct 23, 2023

Contact name

Michael Brent

E-mail(s)

brent@wustl.edu

Phone

314-660-2205

Organization name

Washington University