|
| Status |
Public on Mar 12, 2019 |
| Title |
Nano_OHC_007 |
| Sample type |
SRA |
| |
|
| Source name |
Cochlea
|
| Organism |
Mus musculus |
| Characteristics |
age: p15
background: C3HeB/FeJ
cell type: OHC
|
| Extracted molecule |
polyA RNA |
| Extraction protocol |
Individual cells were dissociated from mouse cochlear tissue using a collagenase digestion and gentle tituration. Cells were then isolated using pulled glass micropipettes, washed to remove extracellular contaminants and re-isolated.
Single cell reverse transciption and PCR amplification was performed using Smartseq2
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
MinION |
| |
|
| Description |
Single-Cell SmartSeq2 Nanopore
|
| Data processing |
For Nanopore data processing, begin with the .fastq file for each cell output by the Nanopore MinION run. Perform Demultiplexing (samples uploaded here have already been demultiplexed) and Alignment using Mandalorion v0.2 (alignment within the mandalorion package is run using gmap). Four cells are multiplexed within each nanopore output .fastq. Demultiplex and align each MinION run output .fastq individually because indexes are repeated between runs.
Perform isoform quantification with Mandalorion Define and Quantify Isoforms.
Mandalorion documentation: https://github.com/christopher-vollmers/Mandalorion and
Mandalorion supporting publication: https://www.nature.com/articles/ncomms16027?WT.feed_name=subjects_adaptive-immunity
Our corresponding publication discusses the use of Nanopore long read sequencing to resolve full length messenger RNA transcripts. Nanopore generated .fastq format files are the raw input data and the output of the bioinformatics analysis are .psl alignment files and corresponding .fasta sequence files.
Genome_build: mm10
Supplementary_files_format_and_content: Output alignment files for individual nanopore reads are in .psl format with corresponding .fasta sequences. The .psl file format is described here https://useast.ensembl.org/info/website/upload/psl.html. .psl format alignment files can be visualized in a genome browser. We prefer to use the IGV browser.
|
| |
|
| Submission date |
May 08, 2018 |
| Last update date |
Mar 12, 2019 |
| Contact name |
Paul T Ranum |
| E-mail(s) |
ranump@chop.edu
|
| Phone |
1 (651) 398-6455
|
| Organization name |
The Children's Hospital of Philadelphia
|
| Department |
Center for Cellular and Molecular Therapy
|
| Lab |
Davidson Lab
|
| Street address |
3501 Civic Center Blvd.
|
| City |
Philadelphia |
| State/province |
PA |
| ZIP/Postal code |
19104 |
| Country |
USA |
| |
|
| Platform ID |
GPL24973 |
| Series (1) |
| GSE114157 |
Insights into the Biology of Hearing and Deafness Revealed by Single-Cell RNA Sequencing |
|
| Relations |
| BioSample |
SAMN09090124 |
| SRA |
SRX4090886 |
