|
| Status |
Public on Jan 30, 2024 |
| Title |
RIP_GPATCH4-His-Flag_A |
| Sample type |
SRA |
| |
|
| Source name |
Flp-In T-Rex HEK293
|
| Organism |
Homo sapiens |
| Characteristics |
cell line: Flp-In T-Rex HEK293
genotype: WT
|
| Treatment protocol |
Expression of tagged proteins was induced by addition of tetracycline for 24 h before crosslinking. Cells were UV crosslinked using light at 254 nm as previously described (Sloan et al., 2015; Memet et al., 2017).
|
| Growth protocol |
HEK293 Flp-In T-Rex cell lines expressing GPATCH4-His-FLAG or the His-FLAG tag alone were grown at 37°C in 5% CO2 in high glucose DMEM supplemented with 10% foetal calf serum .
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Crosslinked RNA-protein complexes were purified and RNAs associated with His-FLAG-tagged proteins were recovered.
Small RNAs <200 nt were enriched and libraries were constructed using the QIAseq miRNA Library Kit Catalog no.331502 according to the maufacturer's instructions.
|
| |
|
| Library strategy |
RIP-Seq |
| Library source |
transcriptomic |
| Library selection |
other |
| Instrument model |
Illumina HiSeq 4000 |
| |
|
| Data processing |
3' Adaptor seuquences (AACTGTAGGCACCATCAAT) were Clipped using Flexbar (version 3.5.0) with the following arguments -qf i1.8 -qt 13 -u 0 -m 15 -n 8 -at RIGHT -z GZ -o -ao 2 -ae 0.1 -as AACTGTAGGCACCATCAAT
Clipped reads were aligned to the human genome using STAR (Version 2.7.10a) with the following arguments --alignEndsType EndToEnd --outFilterMismatchNmax 10 --outFilterMultimapScoreRange 10 --quantMode TranscriptomeSAM GeneCounts --outReadsUnmapped Fastx --outSAMtype BAM SortedByCoordinate --outFilterMultimapNmax 30 --outSAMunmapped Within --outFilterScoreMinOverLread 0 --outFilterMatchNminOverLread 0 --outFilterMatchNmin 16 --alignSJDBoverhangMin 1000 --alignIntronMax 1 --outWigType wiggle --outWigStrand Stranded --outWigNorm RPM --readFilesCommand zcat
Bedgraphs were generated using bedtools (version 2.27.1) with the following arguments genomecov -ibam .bg
Assembly: Homo_sapiens.GRCh38.108
Supplementary files format and content: Aligned reads in bedgraph format
|
| |
|
| Submission date |
Apr 24, 2023 |
| Last update date |
Jan 30, 2024 |
| Contact name |
Katherine Bohnsack |
| E-mail(s) |
katherine.bohnsack@med.uni-goettingen.de
|
| Organization name |
University Medical Centre Göttingen
|
| Street address |
Humboldtallee 23
|
| City |
Göttingen |
| ZIP/Postal code |
37073 |
| Country |
Germany |
| |
|
| Platform ID |
GPL20301 |
| Series (2) |
| GSE230432 |
GPATCH4 regulates rRNA and snRNA 2’-O-methylation in both DHX15-dependent and -independent manners [small RNA-seq] |
| GSE230433 |
GPATCH4 regulates rRNA and snRNA 2'-O-methylation in both DHX15-dependent and -independent manners |
|
| Relations |
| BioSample |
SAMN34353378 |
| SRA |
SRX20083834 |
