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| Status |
Public on Jan 03, 2024 |
| Title |
[ATAC-Seq] shNC_2 |
| Sample type |
SRA |
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| Source name |
SH-SY5Y
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| Organism |
Homo sapiens |
| Characteristics |
cell line: SH-SY5Y
cell type: Neuroblastoma
genotype: control
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| Treatment protocol |
shRNA oligos (shNC and shTDP-43) synthesized from Integrated DNA Technologies (IDT) were annealed and inserted into linearized lentiviral vector pLVX-U6-shRNA1 (Genprice, #PVT2302) using BamH I and EcoR I. The shRNA constructs were then used for lentivirus production. 293T packaging cells were seeded at 5x106 cells per 10 cm tissue culture plate the day before transfection. 1.3 pmol of psPAX2 (Addgene, #12260), 0.72 pmol of pMD2.G (Addgene, #12259), and 1.64 pmol of lentiviral plasmid were co-transfected into 293T cells using polyethylenimine MW 25000 (PEI 25K, Polyscience, #23966-100) at a ratio of 1:3 (w:w). Virus-containing medium was harvested 48 and 72 hr post-transfection and passed through a 0.45 μm PES filter (VWR, #76211-008). The virus was then concentrated 100-fold with Lenti-X concentrator (TaKaRa, #631231). Virus infection was performed by adding 20 μL of lentivirus to ~800,000 SH-SY5Y cells (MOI: 1:3) in a 6-well plate. TDP-43 stable knockdown cells were selected by Puromycin at a concentration of 2 μg/mL for 2 weeks and maintained in medium with 1 μg/mL of Puromycin for long-term use.
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| Growth protocol |
Human neuroblastoma cells SH-SY5Y (ATCC) were purchased from ATCC and cultured in Eagle's Minimum Essential Medium (EMEM) (Corning) and Ham’s F12 medium (Corning) (1:1, v:v) supplemented with 15% v/v fetal bovine serum (FBS) at 37℃ and 5% carbon dioxide (CO2).
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| Extracted molecule |
genomic DNA |
| Extraction protocol |
5x104-1x105 cells were used for ATAC-seq following the manufacturer's instructions (Active Motif, 53150). Cells were lysed with 100 μL of ATAC Lysis Buffer. Accessible DNA region in nuclei was then probed with Tn5 Transposase that inserts sequencing adapters into open regions. The tagged DNA was cleaved, purified, PCR-amplified and sequenced.
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| Library strategy |
ATAC-seq |
| Library source |
genomic |
| Library selection |
other |
| Instrument model |
Illumina HiSeq 2500 |
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| Data processing |
DRIP-seq, hMe-Seal, ATAC-seq reads were first aligned to human genome sequence (hg38) by Bowtie 2 version 2.4.4 with default parameter.
Aligned reads in bam file were sorted by genomic coordinate by samtools.
ATAC-seq reads in each TE were counted and normalized, TE with average reads over 5 in either shNC and shTDP43 were considered as activated TE and were reported
Assembly: hg38
Supplementary files format and content: tab delimated file containing normalized ATAC-seq reads in each activated TE, average reads count in shNC and shTDP43 and logFC
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| Submission date |
Apr 24, 2023 |
| Last update date |
Jan 03, 2024 |
| Contact name |
Bing Yao |
| E-mail(s) |
bing.yao@emory.edu
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| Phone |
3523596473
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| Organization name |
Emory University
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| Department |
Department of Human Genetics
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| Street address |
615 Michael Street
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| City |
Atlanta |
| ZIP/Postal code |
30322 |
| Country |
USA |
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| Platform ID |
GPL16791 |
| Series (2) |
| GSE230406 |
TDP-43 stable deficiency leads to transcriptional dysregulation and transposable elements activation via global R-loops and 5hmC landscape alteration [ATAC-Seq] |
| GSE230410 |
TDP-43 stable deficiency leads to transcriptional dysregulation and transposable elements activation via global R-loops and 5hmC landscape alteration |
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| Relations |
| BioSample |
SAMN34353166 |
| SRA |
SRX20083057 |
| Supplementary data files not provided |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
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