|
| Status |
Public on Sep 04, 2018 |
| Title |
lna_cell_mousehuman_original |
| Sample type |
SRA |
| |
|
| Source name |
1:1 mix of NIH3T3 cell line and 293T cell line
|
| Organisms |
Homo sapiens; Mus musculus |
| Characteristics |
sample type: 1:1 mix of NIH3T3 cell line and 293T cell line
|
| Extracted molecule |
total RNA |
| Extraction protocol |
293FT and NIH3T3 cells were trpsinized and resuspended in a 1:1 mix and captured on the 10x genomics single cell R-seq platform.
10x genomics 3' end single cell RNA-seq kit. Read 1 positions 1-16 contain the 16bp cell barcode, and positions 17-26 contain a 10bp UMI. Read 2 contains the cDNA.
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina HiSeq 4000 |
| |
|
| Description |
Original single cell RNA-seq library from the 10x genomics 3' end library prep. A 1:1 mixture of NIH3T3 cells and 293T cells were isolated
lna_cell_mousehuman_bcs.txt
|
| Data processing |
Data analysis pipeline and custom scripts are provided in a github repository (https://github.com/rnabioco/scrna-subsets). Single cell RNA-Seq libraries were preprocessed to append the read 1 sequence to the paired read 2 read id (append_read_name) followed by quality trimming and poly(A) tail removal from read 2 using cutadapt (v. 1.8.3 -a 'A{{100}}' -O 6 -m 20 -q 10) (Martin 2011). Reads were next aligned with STAR (Dobin et al. 2013) to either the human genome assembly (hg38) for the PBMC experiments or a genome with both human (hg38) and mouse (mm38) sequences for all other experiments. Sequence headers in the human/mouse combined genome were prefixed with either an “H_” or “M_” to designate human or mouse references respectively. Following alignment, BAM files were processed to extract the cell barcode and UMI sequences into tags (CN and BX) within the BAM file (barcode_tag_bam). The cell barcode was error corrected against a list of cell barcodes, either as known well barcodes (Wafergen experiments), or generated from the original 10x genomics single cell libraries processed with the 10x genomics software cellranger (v. 2.1.1). Cell barcodes within edit distance of 1 of the known barcodes were considered valid cell barcodes and corrected to the known barcode. Alignments not designated as multi-mapping that overlapped distinct exonic features were tagged in the BAM file (subread v. 1.6.0) (Liao et al. 2014). Gencode v25 annotations was used for human data, and a union reference containing Gencode v25 and mouse v11 was used for human/mouse datasets. UMIs per gene were enumerated per cell using umi-tools (v 0.5.3) (Smith et al. 2017) using the directional method to disambiguate similar UMI sequences.
Genome_build: mm38, hg38
Supplementary_files_format_and_content: Text files containing cell barcode sequences, count matrices of UMIs per gene per cell are provided. For the TCR single cell libraries JSON files are provided with detailed annotations for each assembled contig per cell.
|
| |
|
| Submission date |
Sep 04, 2018 |
| Last update date |
Sep 05, 2018 |
| Contact name |
Kent Augustus Riemondy |
| Organization name |
University of Colorado School of Medicine
|
| Department |
Biochemistry and Molecular Genetics
|
| Street address |
12801 E 17th Ave
|
| City |
Aurora |
| State/province |
Colorado |
| ZIP/Postal code |
80045 |
| Country |
USA |
| |
|
| Platform ID |
GPL25431 |
| Series (1) |
| GSE119428 |
Recovery and analysis of transcriptome subsets from pooled single-cell RNA-seq libraries |
|
| Relations |
| BioSample |
SAMN09951391 |
| SRA |
SRX4637901 |
