|
| Status |
Public on Jul 15, 2022 |
| Title |
HeLa parental cell line rep 1 |
| Sample type |
SRA |
| |
|
| Source name |
HeLa
|
| Organism |
Homo sapiens |
| Characteristics |
cell type: Cervical adenocarcinoma
strain: HeLa-S3
genotype: wild type
|
| Growth protocol |
HeLa S3 cells were cultured in DMEM supplemted with 10% Fetal Bovine Serum and Penicillin-Streptomycin at 37°C in 5% CO2.
|
| Extracted molecule |
polyA RNA |
| Extraction protocol |
Total RNA was extracted with Trizol reagent (Ambion), treated with Ambion™ DNase I (RNase-free) extracted with phenol/chloroform. RNA samples were quantified and quality tested by Agilent 2100 Bioanalyzer RNA assay (Agilent technologies, Santa Clara, CA).
RNA libraries were prepared for Oxford Nanopore MinION sequencing using PCR-cDNA Barcoding (SQK-PCB109) protocol.
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
MinION |
| |
|
| Data processing |
Primary data acquisition was performed by MinKNOW.
Basecalling and demultiplexing of Fast5 files obtained by sequencing were performed using Guppy v4.2.2.
Reads with Q-score >7 were mapped to hg38 genome using Minimap2 v2.17 with parameters -ax splice.
To assemble and quantify transcripts from Nanopore reads we used FLAIR v1.4. BAM files obtained with Minimap2 were converted into BED12 files using the bam2Bed12.py utility included in FLAIR. After this, splice junctions of reads were corrected using FLAIR correct module using both the GENCODE v33 GTF file and short Illumina reads of the corresponding samples. BED12 files were then filtered to maintain only reads with at least one splice junction.
Filtered BED12 files from all samples were used to assemble the transcriptome using the FLAIR collapse module, providing the GENCODE GTF file to annotate the results and requiring at least 10 supporting reads to identify novel transcripts. Transcripts from both the GENCODE and FLAIR GTF files were quantified using the FLAIR quantify module.
Genome_build: hg38
Supplementary_files_format_and_content: GTF file describing the transcriptome assembled by FLAIR.
Supplementary_files_format_and_content: Tab-delimited text file including TPM values in each sample calculated for each transcript assembled by FLAIR.
|
| |
|
| Submission date |
Nov 18, 2021 |
| Last update date |
Jul 15, 2022 |
| Contact name |
Grazia Daniela Raffa |
| E-mail(s) |
graziadaniela.raffa@uniroma1.it
|
| Organization name |
Sapienza Università di Roma
|
| Street address |
P.le Aldo Moro 5
|
| City |
Rome |
| ZIP/Postal code |
00185 |
| Country |
Italy |
| |
|
| Platform ID |
GPL24106 |
| Series (1) |
| GSE189138 |
TGS1 controls snRNA 3' end processing, prevents neurodegeneration and ameliorates SMN-dependent neurological phenotypes in vivo (Nanopore) |
|
| Relations |
| BioSample |
SAMN23284652 |
| SRA |
SRX13167050 |
