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| Status |
Public on Jun 10, 2021 |
| Title |
95% HEK293T [ZZ203-PP04-A03] |
| Sample type |
SRA |
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| Source name |
HEK293T/CCRM-CEM mixture
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| Organism |
Homo sapiens |
| Characteristics |
x: ZZ203-PP04-A03
sample_id: ZZ203-PP04-A03
panel: broad
construct: directPCR
instrument_type: NEXTSEQ
expected_neg: 0
ngs_repset: 291
ngs_repnum: 1
ngs_prep: ZZ203
pcr_primer_plate: PP04
pcr_primer_well: A03
read_pattern: 2x38
pcr_plate_ignore: 2
pcr_well_ignore: A03
seq_instrument: nwgc-nextseq-20190617
index_1: i7291
barcode_1: GAACGCGC
index_2: i5291
barcode_2: GAGATCAT
pcr_plate: ZZ203-PCR2
pcr_well: A03
primer_plate: PP04
primer_well: A03
dsp_date: 20190607
dsp_instrument: Beta7
slide_type: 293T CPA
dsp_plate_id: Beta 7: 1011220607190
dsp_aoi_by_plate: 3
dsp_well: A03
dsp_aoi_by_slide: 3
aoi_size: 300
aoi_area: 70686
exp_repset: 99
exp_repnum: 2
dsp_repset: 140
dsp_repnum: 1
segment_type: solid
sample_name: HEK293T--CCRF-CEM
sample_type: mixed cell line
notes: 95% HEK293T
uniq_id: ZZ203-PP04-A03
hk_geo: 1004.081466
q3: 173.389876
neg_geomean_bkp: 28.07527477
neg_geomean_lkp: 17.55553075
hk_factor: 3.232847915
q3_factor: 2.661005394
neg_bkp_factor: 2.151754806
neg_lkp_factor: 2.317425352
area_pixels: 446469
mean_intensity_blue: 2398
mean_intensity_green: 326
mean_intensity_yellow: na
mean_intensity_red: 440
mix1: HEK293T
mix2: CCRF-CEM
mixprop: 0.95
hkfactor: 7.92568374110767
bgmean: 4.31594911147382
bg.lkp: 3.8202404502754
bg.bkp: 4.81165777267223
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| 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.
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| Library strategy |
OTHER |
| Library source |
transcriptomic |
| Library selection |
other |
| Instrument model |
NextSeq 550 |
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| Description |
photocleaved oligonucleotide barcodes
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
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| 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
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.
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| Submission date |
May 20, 2021 |
| Last update date |
Jun 10, 2021 |
| Contact name |
Patrick Danaher |
| Organization name |
NanoString Technologies, Inc.
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| Street address |
530 Fairview Ave N
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| City |
Seattle |
| State/province |
WA |
| ZIP/Postal code |
98109 |
| Country |
USA |
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| Platform ID |
GPL21697 |
| Series (2) |
| GSE174746 |
GeoMx Cancer Transcriptome Atlas (CTA) mRNA assay on an FFPE cell pellet array of mixed HEK293T and CCRF-CEM cell lines. |
| GSE175927 |
GeoMx Cancer Transcriptome Atlas (CTA) mRNA assay |
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| Relations |
| BioSample |
SAMN19284616 |
| SRA |
SRX10947777 |
| Supplementary data files not provided |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data are available on Series record |
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