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| Status |
Public on Dec 28, 2018 |
| Title |
APTX-POS-2 |
| Sample type |
SRA |
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| Source name |
sarcoma cell line
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| Organism |
Homo sapiens |
| Characteristics |
cell line: U2OS
genotype: APTX over expression
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| Treatment protocol |
no treatment
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| Growth protocol |
The cells were in culture for two-three weeks before RNA extraction. U2OS cells were grown at a density of 2–3 × 106 cells into a T75 flask, in DMEM medium (Life Technologies) supplemented with 10% FBS (Sigma) and 1% penicillin-streptomycin. AOA1 patients cells were cultured in RPMI 1640 medium (Life Technologies) supplmented with 10% FBS and 1% penicillin-streptomycin. The cells were cultured at a density of 5-10 x 106 in a T75 flask.
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| Extracted molecule |
total RNA |
| Extraction protocol |
total RNA was extracted with RNeasy Mini Kit (Qiagen).
For library construction, polyA-containing mRNA molecules were isolated with magnetic beads. Following purification, the mRNA was fragmented into small pieces. The cleaved RNA fragments were copied into first strand cDNA using reverse transcriptase and random primers. This is followed by second strand cDNA synthesis using DNA Polymerase I and RNase H. These cDNA fragments then have the addition of a single 'A' base and subsequent ligation of the adapter. The products are then purified and enriched with PCR amplification. We then quantified the PCR yield by Qubit and pooled samples together to make a single strand DNA circle (ssDNA circle), which gave the final library.
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| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
BGISEQ-500 |
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| Description |
APTX-KO U2OS cells expressing GFP-APTX
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| Data processing |
Base calling software: ZebraCall, V2
SOAPnuke for filter reads: version:v1.5.2 parameters: -l 15 -q 0.5 -n 0.1 website: https://github.com/BGI-flexlab/SOAPnuke
HISAT (Hierarchical Indexing for Spliced Alignment of Transcripts) for mapping step. HISAT2: Figure3 HISATmappingdemoshow. Version: v2.0.4 Parameters: --phred64 --sensitive --no-discordant --no-mixed -I 1 -X 1000 Website: http://0-www-ccb-jhu-edu.brum.beds.ac.uk/software/hisat
StringTie to reconstruct transcripts, and use Cuffcompare(Cufflinks tools) to compare reconstructed transcripts to reference annotation. CPC to predict coding potential of novel transcripts StringTie: Version: v1.0.4 Parameters: -f 0.3 -j 3 -c 5 -g 100 -s 10000 -p 8 Website: http://0-ccb-jhu-edu.brum.beds.ac.uk/software/stringtie Cufflinks: Version: v2.2.1 Parameters: -p 12 Website: http://cole-trapnell-lab.github.io/cufflinks CPC: Version: v0.9-r2 Parameters: Default Website: http://cpc.cbi.pku.edu.cn
rMATS to detect differentially splicing gene. rMATS: Version: v3.2.5 Parameters: -analysis U -t paired -a 8 Website: http://rnaseq-mats.sourceforge.net
Bowtie2 for mapping reads to reference, RSEM for calculating gene expression level. Bowtie2 : Version: v2.2.5 Parameters: -q --phred64 --sensitive --dpad 0 --gbar 99999999 --mp 1,1 --np 1 --score-min L,0,-0.1 -I 1 -X 1000 --no- mixed--no-discordant -p1-k200 Website: http://bowtie-bio.sourceforge.net/ Bowtie2 /index.shtml RSEM : Version: v1.2.12 Parameters: default Website: http://deweylab.biostat.wisc.edu/ RSEM
Genome_build: hg19
Supplementary_files_format_and_content: excel files include RPKM values for each Sample
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| Submission date |
Dec 27, 2018 |
| Last update date |
Dec 31, 2018 |
| Contact name |
Jin Zheng |
| E-mail(s) |
zhj821107@icloud.com
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| Phone |
+4550245870
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| Organization name |
University of Copenhagen
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| Street address |
3b Blegdamsvej
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| City |
Copenhagen |
| ZIP/Postal code |
2200 |
| Country |
Denmark |
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| Platform ID |
GPL23227 |
| Series (1) |
| GSE124412 |
Diminished OPA1 expression and impaired mitochondrial morphology and homeostasis in Aprataxin-deficient cells |
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| Relations |
| BioSample |
SAMN10654221 |
| SRA |
SRX5184299 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM3532174_APTX-Pos-2_gene_fpkm.xls.gz |
2.8 Mb |
(ftp)(http) |
XLS |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
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