|
| Status |
Public on May 31, 2017 |
| Title |
Control RNAi batch 6 |
| Sample type |
SRA |
| |
|
| Source name |
luci_2
|
| Organism |
Homo sapiens |
| Characteristics |
cell line: HeLa Kyoto cells
treatment: control siRNA
experimental batch: 6
|
| Treatment protocol |
2 x 10^5 HeLa Kyoto cells were seeded into 6-well plates 24 h prior to transfection to reach a confluence of 30 – 40%. Small interfering RNA (siRNA) stock solutions (100 µmol in 1xUniversal Buffer) were diluted 1:5 in sterile water. 4 µl Oligofectamine transfection reagent was diluted in 11 µl Opti-MEM and incubated at RT for 5 min. Meanwhile, 10 µl of siRNA dilution was mixed with 175 µl Opti-MEM. 15 µl transfection reagent dilution was added and incubated for 20 min at RT. Cells were washed twice with sterile PBS, growth medium replaced by 800 µl DMEM not containing FCS or antibiotics and the transfection mix added drop wise. After 4 h at 37°C in the incubator 500 µl DMEM supplemented with 30% FCS was added. Cells were harvested and analyzed 2 to 3 days after transfection.
|
| Growth protocol |
HeLa Kyoto cells were grown in DMEM medium (PPA), supplemented with 10% FCS (Sigma-Aldrich) and 1% penicillin / streptomycin (PAA) at 37°C and 5% CO2
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA from HeLaK cells subjected to PWWP2A siRNAs and control siRNAs was isolated utilizing the RNeasy mini kit (Qiagen) according to manufacturer’s instructions. The quality of the RNA (amount and integrity) was assessed using the BioAnalyzer and a RNA 6000 pico kit (Agilent). mRNA isolation, fragmentation, first and second strand cDNA synthesis and end repair were carried out using the NEBNext Ultra RNA library prep kit for Illumina (NEB) following the manufacturer’s instructions
cDNA sequencing libraries were established with the MicroPlex Library Preparation Kit (Diagenode) as described above
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina HiSeq 1500 |
| |
|
| Data processing |
RNA-seq reads were aligned to the GRCh38 genome assembly using STAR version 2.4.1c
Gene-level expression quantitation was performed using rsem version 1.2.22
Genome_build: GRCh38
Supplementary_files_format_and_content: tab-delimited text file for each sample with gene id, transcript ids, length, effective length, expected count, TPM, FPKM
|
| |
|
| Submission date |
Feb 17, 2016 |
| Last update date |
May 15, 2019 |
| Contact name |
Tobias Straub |
| E-mail(s) |
tstraub@med.uni-muenchen.de
|
| Organization name |
LMU Munich
|
| Department |
Biomedical Center, Bioinformatics
|
| Street address |
Großhadener Str. 9
|
| City |
Martinsried |
| ZIP/Postal code |
82152 |
| Country |
Germany |
| |
|
| Platform ID |
GPL18460 |
| Series (2) |
| GSE78003 |
Multivalent binding of PWWP2A to H2A.Z-marked transcriptional active chromatin regulates mitosis and organ development [RNA-seq] |
| GSE78009 |
Multivalent binding of PWWP2A to H2A.Z-marked transcriptional active chromatin regulates mitosis and organ development |
|
| Relations |
| SRA |
SRX1589777 |
| BioSample |
SAMN04503794 |
