|
| Status |
Public on Nov 14, 2020 |
| Title |
A20-102 A40A20-102 A40_ROI027PanCK+ |
| Sample type |
SRA |
| |
|
| Source name |
FFPE Rapid Autopsy Tissue
|
| Organisms |
Homo sapiens; Severe acute respiratory syndrome coronavirus 2 |
| Characteristics |
case number: Case 7
patient id: A20-102 A40
roi number: 27
segment type: PanCK_Pos
tissue substructure: Alveoli
sars-cov-2 rna ish: Negative
patient viral load: 1.30E-02
roi x coordinate: 82.267
roi y coordinate: -27.331
rawreads: 471231
trimmedreads: 470367
stitchedreads: 452393
alignedreads: 449590
deduplicatedreads: 222902
sequencingsaturation: 50.4210502902645
note: NA
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Samples were incubated with DNA-oligo barcoded RNA-ISH probes which were conjugated with a UV-photocleavable linker following standard ISH protocols, along with flourescently labeled antibodies for visualization of morphological structures. Regions of interest within the tissue were illuminated with UV light and oligo barcodes were physically aspirated from the tissue and collected into microtiter plates by the GeoMx® Digital Spatial Profiler (DSP) platform. For more information about DSP protocols please see Merritt et al. Nature Biotech 2020 (doi: 10.1038/s41598-020-63539-x)
Each collection of oligo tags from one well (representing an AOI from the tissue section) was indexed with i7xi5 unique dual indexes using GeoMx SeqCode primers with 18 cycles of PCR. After PCR, indexed AOIs were pooled and purified in two rounds of AMPure XP PCR purification using 1.2x bead:sample ratio.
|
| |
|
| Library strategy |
OTHER |
| Library source |
transcriptomic |
| Library selection |
other |
| Instrument model |
NextSeq 550 |
| |
|
| Description |
Samples were pooled during library preparation and split across lanes. Individual FASTQ files were generated for each lane, forward & reverse primers. The Fastq files contain the same identifier as the sample name for aggregation
DSP-1001250365000-D02.dcc
|
| Data processing |
Library strategy: GeoMx Seq
Adapter trimming with trim galore, trimGaloreOpts = " --hardtrim5 26 --dont_gzip"
If paired-end, merge overlapping R1 and R2 with flash2, flash2Opts = " -m 26 -e 26 -f 26 -s 1 -r 27"
Extract UMIs in bowtie2, umiExtractOpts = " --bc-pattern=NNNNNNNNNNNNNN"
Align RTS_IDs (probe barcodes) using bowtie2, bowtie2Opts = " --end-to-end -L 4 --trim5 0 --trim3 0 --norc"
Deduplication using UMI-tools, umiDedupOpts = " --edit-distance-threshold=1"
Genome_build: N/A, sequencing of synthetic tags and alignment to whitelist of RTS_IDs (probe barcodes) found in the config.ini output from the GeoMx DSP platform, see attached NanoString GeoMx COVID19 Immune Response Atlas v1 Platform file & GeoMx_COVID19_v1.0.pkc file
Supplementary_files_format_and_content: Digital Count Conversion (DCC) file format outputted from GeoMx NGS Pipeline, contains software versions, scan attributes, GeoMx NGS pipeline parameters and output metrics, Q30 scores, and list of deduplicated counts per RTS_ID (probe barcode)
Supplementary_files_format_and_content: Values represented in the collapsed target counts tab are the geometric mean of the probes for a given, removing any targets flagged as outliers. Analyzed counts represent the upper quartile normalized collapsed counts across the study after removing QC flagged segments.
|
| |
|
| Submission date |
Nov 13, 2020 |
| Last update date |
Nov 15, 2020 |
| Contact name |
Jason W Reeves |
| E-mail(s) |
jreeves@nanostring.com
|
| Organization name |
Nanostring
|
| Department |
R&D
|
| Street address |
530 Fairview Blvd
|
| City |
Seattle |
| State/province |
WA |
| ZIP/Postal code |
98109 |
| Country |
USA |
| |
|
| Platform ID |
GPL29228 |
| Series (2) |
| GSE159787 |
Temporal and Spatial Heterogeneity of Host Response to SARS-CoV-2 Pulmonary Infection [gene expression] |
| GSE159788 |
Temporal and Spatial Heterogeneity of Host Response to SARS-CoV-2 Pulmonary Infection |
|
| Relations |
| BioSample |
SAMN16792974 |
| SRA |
SRX9504800 |
