|
| Status |
Public on Jan 11, 2021 |
| Title |
MouseSpermONT |
| Sample type |
SRA |
| |
|
| Source name |
sperm
|
| Organism |
Mus musculus |
| Characteristics |
strain: C57B6
age: adult
tissue/cell type: cauda sperm
molecule subtype: full length RNA
|
| Treatment protocol |
NA
|
| Growth protocol |
Mouse sperm samples were acquired from 10-week until 6-month adult male mice. Mice were housed in a temperature (64-79°F) and humidity (30-70%) controlled room with 12 light/12 dark cycle.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Before RNA purification, sperm cells were lysed by 1% SDS and 0.1M DTT, with 10 min incubatin on 37 degree. Sperm RNAs were isolated using Trizol followed by acid phenol extraction. rRNA depletion was done for Illumina RNAseq.
PacBio SMRTseq uses PacBio library construction protocol. Illumina sequencing uses standard RNAseq library construction protocol. Nanopore library was constructed using cDNA-PCR Sequencing Kit (SQK-PCS109, ONT)
PacBio, Illumina RNAseq and Oxford Nanopore Technologies
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
PromethION |
| |
|
| Data processing |
LoRDEC (version 0.5.3, parameters -k 17 -s 3 -a 50000) was used to perform error correction on the long reads (LRs) with short reads (SRs) as input.
The corrected LRs were aligned to the mm10 or hg38 reference genome using GMAP (version 2014-12-24, parameters -t 10 -B 5 -A --nofails -f samse -n1). The aligned LRs with clipping region longer than 100-nt at either end were filtered out.
The SRs were aligned to the mm10 or hg38 reference genome using HISAT2 (version 2.0.4, parameters --ignore-quals --dta --threads 10 --max-intronlen 150000).
Illumina short reads were quantified using Salmon (v0.8.2), and TPM values were summaized in the processed data table. Sperm samples were further normalized to the renilla spikein number.
SpermPacBio1-6 were combined, and TestisPacBio1-5 were combined.
ONT sequencing reads were aligned to the mouse mm10 reference genome using minimap2 (version 2.11-r797) with parameter "-x splice -a".
Genome_build: mm10, hg38
Supplementary_files_format_and_content: Long reads numbers mapped to each transcript have been summarized into Summary.xlsx table. Transcript TPM values were also summarized in the table.
|
| |
|
| Submission date |
Jan 11, 2021 |
| Last update date |
Jan 13, 2021 |
| Contact name |
Xin Li |
| E-mail(s) |
xin_li@urmc.rochester.edu
|
| Organization name |
University of Rochester Med Center
|
| Department |
Biochemistry and biophysics
|
| Lab |
Xin Li
|
| Street address |
601 Elmwood Ave.
|
| City |
Rochester |
| State/province |
NY |
| ZIP/Postal code |
14642 |
| Country |
USA |
| |
|
| Platform ID |
GPL26624 |
| Series (1) |
| GSE137490 |
Single-molecule long-read sequencing reveals a conserved intact long RNA profile in sperm |
|
| Relations |
| BioSample |
SAMN17284305 |
| SRA |
SRX9822381 |
