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| Status |
Public on Jul 10, 2024 |
| Title |
RNASEQ, clone SC9, NELFB, replicate 1 |
| Sample type |
SRA |
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| Source name |
HAP1 dCAS9-KRAB cSC9
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| Organism |
Homo sapiens |
| Characteristics |
cell line: HAP1 dCAS9-KRAB cSC9
cell type: HAP1
genotype: NELFB Knockdown
antibody: NA
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| Treatment protocol |
The cell line was infected with lentiviral vectors (Lentiguide Puro: Addgene #52963) containing guide RNA for: ADAM22 (negative control), NELFE or NELFB. We produced lentiviral particles by transfecting HEK293T with Calcium Phosphate transfection and using pREV, pMDL, and pVSV as packaging vec-tors. Supernatants from a 10 cm dish were collected 48 hours after transfection, filtered, and concentrated to 250 µl using Amicon filters, aliquoted, and stored at -80°C. HAP1 cells were infected with 30 µl of concentrated virus and then selected for puromycin resistance 48 hours after transduction. We performed mRNA extraction 144 hours following lentiguide transduction for RNAseq
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| Growth protocol |
HAP1 cell line were grown in IMDM (Gibco, 15070063) FBS0 10%, 1% penicillin-streptomycin (Gibco, 15070063) . Cells were passaged every 2 days. Mycoplasma contamination was ruled out by regular testings (#LT07-318; Lonza).
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| Extracted molecule |
polyA RNA |
| Extraction protocol |
One million cells were harvested, washed once in cold PBS, resuspended in 600 µL of RTL buffer (RNeasy Mini kit, 74104, Qiagen) and stored at -80°C until subsequent RNA isolation. RNA was isolated using the RNeasy Mini kit (74104, Qiagen).
Libraries were prepared using the TruSeq® RNA LT kit and TruSeq RNA Single Indexes (Illumina). Libraries were sequenced with single-end 65-bp reads on a HiSeq 2500 platform
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| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina HiSeq 2500 |
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| Data processing |
RNA-seq reads were subjected to quality control using FastQC v0.11.6. Reads were aligned to the human reference genome (GRCh38, GRCh38_no_alt_analysis_set_GCA_000001405.15;https://www.encodeproject.org/files/GRCh38_no_alt_analysis_set_GCA_000001405.15/) using STAR 2.5.4a (Dobin et al. 2013) with parameters --clip5pNbases 0 --outWigStrand Unstranded. Gene-level count tables were generated while mapping using Gencode v24 primary assembly annotations.
Genome_build: GRCh38
Assembly: hg38
Supplementary files format and content: BigWig files and Rawcounts
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| Submission date |
Mar 19, 2024 |
| Last update date |
Jul 10, 2024 |
| Contact name |
Stefano Giustino Manzo |
| E-mail(s) |
stefano.manzo@unimi.it
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| Phone |
+393283622562
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| Organization name |
University of Milan
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| Department |
Biosciences
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| Street address |
Via Celoria 26, Dipartimento Bioscienze, Torre A, Piano 4
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| City |
Milano |
| State/province |
MI |
| ZIP/Postal code |
20133 |
| Country |
Italy |
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| Platform ID |
GPL16791 |
| Series (2) |
| GSE261953 |
Identification of Chromatin proteins involved in gene repression in lamina-associated domains (RNA-Seq) |
| GSE261955 |
Identification of Chromatin proteins involved in gene repression in lamina-associated domains |
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| Relations |
| BioSample |
SAMN40546973 |
| SRA |
SRX23994716 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM8155093_CRISPrI_NELFB_BR1_UnknownStranded_SE_Signal.Unique.str1.out.bw |
106.3 Mb |
(ftp)(http) |
BW |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
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