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| Status |
Public on Mar 22, 2023 |
| Title |
eaSSc_MT2014 [MT2.014] |
| Sample type |
SRA |
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| Source name |
CD1c+ cDCs
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| Organism |
Homo sapiens |
| Characteristics |
cell type: CD1c+ cDCs
tissue: Peripheral blood
disease state: early systemic sclerosis
location: Milan
seks: Female
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| Extracted molecule |
total RNA |
| Extraction protocol |
Human CD1c+ dendritic cells were purified from heparinised whole blood of SSc patients and matched healthy donors after centrifugation over Ficoll-Paque gradient. Briefly, CD1c+ dendritic cells were purified from PBMCs using the anti-BDCA1 microbeads (Miltenyi Biotec), on the autoMACs Pro Separator (Miltenyi Biotec) according to manufacturer’s protocol. Cells were lysed in RLTplus buffer (Qiagen) containing 1% (v/v) beta-mercaptoethanol (Sigma). Total RNA was purified with the RNeasy Mini Kit (Qiagen), according to the manufacturer’s instructions. DNAse treatment (RNAse Free DNase I set, Qiagen) on column was performed.
RNA-Seq libraries were prepared using the TruSeq Stranded kit (Illumina, San Diego, CA, USA) after poly (A) capture, according to the manufacturer’s instructions. Libraries were sequenced on the HiSeq 2000 system (Illumina), using 100bp paired-end reads.
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| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina HiSeq 2000 |
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| Description |
FCH5WGGBBXX-HKHUMvknEABHRABPEI-211
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| Data processing |
Quality filtering according to the BGI pipeline
Reads were aligned to the GrCh38 reference human genome and the homo sapiens transcriptome (Ensembl, version 79), using the STAR aligner
Summed exon read counts per gene were calculated using the Python package HTSeq, using annotations from the GrCh38 built from the human genome
Between lane normalization using the upper quartile normalization method was performed using the Bioconductor/R package EDAseq
To account for batch effects arising from the inclusion of samples from different geographic locations (The Netherlands and Italy), we applied the generalized linear model from the Bioconductor/R package RUVseq using the RUVr function for k = 2 factors of unwanted variation
Differential expression analysis was performed using the negative binomial distribution-based method implemented in DESeq2 on the normalized summed exon read counts per gene
Normalized gene expression levels were converted variance stabilised data (VSD), calculated according to DESeq2 instructions
Genome_build: GrCh38
Supplementary_files_format_and_content: Tab delimited .txt files include counts and normalized expression values (VSD) for each sample.
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| Submission date |
Oct 20, 2021 |
| Last update date |
Mar 22, 2023 |
| Contact name |
Aridaman Pandit |
| Organization name |
University Medical Center Utrecht
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| Street address |
Heidelberglaan 100
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| City |
Utrecht |
| ZIP/Postal code |
3584 CX |
| Country |
Netherlands |
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| Platform ID |
GPL11154 |
| Series (2) |
| GSE186198 |
Nuclear receptor subfamily 4A signaling as a key disease pathway of CD1c+ dendritic cell dysregulation in systemic sclerosis [RNA-seq] |
| GSE186199 |
Nuclear receptor subfamily 4A signaling as a key disease pathway of CD1c+ dendritic cell dysregulation in systemic sclerosis |
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| Relations |
| BioSample |
SAMN22420539 |
| SRA |
SRX12698929 |
| Supplementary data files not provided |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data are available on Series record |
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