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| Status |
Public on Jan 31, 2021 |
| Title |
0.25 h_Flu_rep2 |
| Sample type |
SRA |
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| Source name |
Candida albicans yeast cells
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| Organism |
Candida albicans |
| Characteristics |
strain: SC5314
treatment: 50 µM fluconazole
timepoint: 0.25 h
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| Treatment protocol |
Cultures were either left untreated or treated with CuSO4 (10 µM), fluconazole (50 µM), or both CuSO4 and fluconazole. Cells were harvested at 0, 0.25, 0.67, 1, 3, 5, and 7 h, flash frozen in liquid nitrogen, and stored at −80 °C prior to processing.
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| Growth protocol |
C. albicans cells were streaked onto a YPD agar plate from a frozen glycerol stock and incubated at 30 °C for 24 h. A single colony was used to inoculate liquid YPD growth medium (10 mL), and this culture was incubated overnight (~18 h) at 30 °C with shaking at 200 rpm. The overnight culture of C. albicans in YPD medium was inoculated into fresh YPD medium at an OD600 of 0.3 and this subculture was allowed to grow for 3 h at 30 °C with shaking at 200 rpm prior to treatment.
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| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was isolated from frozen pellets using a Qiagen RNeasy Mini Kit, and DNase was treated with a Turbo DNA-free kit (Roche) according to the manufacturer’s instructions. RNaseZap RNase Decontamination Solution (Invitrogen) was used on all materials and surfaces to preserve RNA integrity.
RNA-seq libraries were prepared using the commercially available KAPA Stranded mRNA-Seq Kit. In brief, mRNA transcripts were first captured using magnetic oligo-dT beads, fragmented using heat and magnesium, and reverse transcribed using random priming. During the second strand synthesis, the cDNA:RNA hybrid was converted into double-stranded cDNA (dscDNA) and dUTP incorporated into the second cDNA strand, effectively marking the second strand. Illumina sequencing adapters were then ligated to the dscDNA fragments and amplified to produce the final RNA-seq library. The strand marked with dUTP was not amplified, allowing strand-specificity sequencing. Libraries are typically indexed using a dual indexing approach allowing for multiple libraries to be pooled and sequenced on the same sequencing lane on a NovaSeq 6000 Illumina sequencing platform. Before pooling and sequencing, fragment length distribution and library quality were first assessed on a Fragment Analyzer. All libraries were then pooled in equimolar ratio and sequenced.
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| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina NovaSeq 6000 |
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| Data processing |
The raw reads were trimmed by Cutadapt (v 1.12) for quality filtering and adaptor removal.
The paired-reads were aligned to the transcriptome of Candida albicans SC5314 (GCF_000182965.3) by using STAR (v 2.5.3)
After the normalization, the read counts of all genes for each sample were then estimated with RSEM (v2.20).
The gene count data was converted to counts per million (CPM) and normalized by adjusting with the effective library size via the 'calcNormFactors' function implemented in the R package edgeR (v. 3.26.7)
The significant DEGs, comparing to NT condition, were determined by using the negative binomial generalized linear models with quasi-likelihood tests ('glmQLFTest'functions in the edgeR package) with FDR p-values of less than 0.05.
Genome_build: We also carried out GO term and KEGG pathway enrichment for DEGs in each cluster by using Fisher's Exact Test with multiple testing correction of FDR threshold 0.05.
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| Submission date |
Oct 19, 2020 |
| Last update date |
Jan 31, 2021 |
| Contact name |
Katherine J. Franz |
| E-mail(s) |
katherine.franz@duke.edu
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| Phone |
9196601541
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| Organization name |
Duke University
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| Department |
Chemistry
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| Street address |
2103 French Science Center, 124 Science Dr., Box 90346
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| City |
Durham |
| State/province |
NC |
| ZIP/Postal code |
27708-0346 |
| Country |
USA |
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| Platform ID |
GPL28323 |
| Series (1) |
| GSE159545 |
Copper Availability Influences the Transcriptomic Response of C. albicans to Fluconazole Stress |
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| Relations |
| BioSample |
SAMN16475354 |
| SRA |
SRX9296716 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM4832270_S10.genes.results.txt.gz |
107.2 Kb |
(ftp)(http) |
TXT |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data are available on Series record |
| Processed data provided as supplementary file |
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