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Status |
Public on Feb 06, 2019 |
Title |
RNASeq_RMCO3 |
Sample type |
SRA |
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Source name |
Cord blood CD34 cells
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Organism |
Homo sapiens |
Characteristics |
mutation status: RUNX1mut transduced harvest time: NA cell type: Cord blood CD34 cells
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Extracted molecule |
total RNA |
Extraction protocol |
Total RNA was extracted from 1x10e6 iPSC control and RUNX1mut cells using the RNeasy mini kit (Qiagen) and performing on-column DNaseI treatment. Ribosomal RNA was depleted by using the ribo-zero rRNA removal kit (Illumina). RNA was fragmented in 300 bp fragments by incubation for 7.5 minutes in fragmentation buffer (200 mM Tris-acetate, 500 mM Potassium Acetate, 150 mM Magnesium Acetate (pH 8.2)). First strand cDNA synthesis was performed using superscript III (Life Technologies), followed by synthesis of the second cDNA strand. Libraries were generated using the Kapa hyper prep kit (KAPA Biosystems). Libraries were generated using the Kapa hyper prep kit. End repair and A-tailing was performed on the double strand DNA using end repair and A-tailing buffer. Subsequently, the adapters were ligated and a post-ligation cleanup was performed using Agencourt AMPure XP reagent. The libraries were amplified by PCR using the Kapa Hifi hotstart readymix and primer mix, 10 cycles. Samples were purified using the Qiaquick MinElute PCR purification kit and 300 bp fragments were selected from E-gel and the size was checked on the agilent bioanalyzer. Samples were sequenced on the Illumina NextSeq 500.
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Library strategy |
RNA-Seq |
Library source |
transcriptomic |
Library selection |
cDNA |
Instrument model |
Illumina NextSeq 500 |
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Data processing |
For expression analyses, the hg19 reference genome index was first generated using STAR aligner with UCSC gene annotation. Paired-end reads were mapped to the indexed genome in two-pass mode with default parameters, to increase alignment accuracy and sensitivity. Stranded gene-level read counts were enumerated at the same time. Expression quantification for each RefSeq gene was performed by Cuffnorm function in Cufflinks, to estimate Fragments Per Kilobase per Million aligned reads value (FPKM). All alignment files were scaled to RPKM-normalized read coverage files using deepTools for visualization. Genome_build: hg19 Supplementary_files_format_and_content: Gene-level read counts were enumerated by STAR tool. bigWig files were generated using deepTools by scaling read depth to RPKM-normalized coverage
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Submission date |
Mar 15, 2018 |
Last update date |
Nov 11, 2021 |
Contact name |
Joost Martens |
E-mail(s) |
joost.martens@ru.nl
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Phone |
0243780645
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Organization name |
Radboud University
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Department |
RIMLS
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Lab |
Molecular Biology
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Street address |
Geert Grooteplein 28
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City |
Nijmegen |
State/province |
Nederland |
ZIP/Postal code |
6525GA |
Country |
Netherlands |
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Platform ID |
GPL18573 |
Series (2) |
GSE111918 |
RUNX1 mutations lead to a myeloid differentiation block by altering the RUNX1 transcriptional program (RNA-Seq) |
GSE111919 |
RUNX1 mutations lead to a myeloid differentiation block by altering the RUNX1 transcriptional program |
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Relations |
BioSample |
SAMN08720343 |
SRA |
SRX3800489 |
Supplementary file |
Size |
Download |
File type/resource |
GSM3044964_RMCO3_Mylene_CB.bw |
27.8 Mb |
(ftp)(http) |
BW |
GSM3044964_RMCO3_Mylene_CB_ReadsPerGene.out.tab.gz |
139.9 Kb |
(ftp)(http) |
TAB |
SRA Run Selector |
Raw data are available in SRA |
Processed data provided as supplementary file |
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