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| Status |
Public on Mar 30, 2018 |
| Title |
P1_left-midbrain_Scars |
| Sample type |
SRA |
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| Source name |
left midbrain single cells
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| Organism |
Danio rerio |
| Characteristics |
cas9 injection: protein
FISH id: P1
tissue: left midbrain
sorted plates: 1
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| Extracted molecule |
total RNA |
| Extraction protocol |
After organ isolation, live single cells are sorted into 384 well plates containing mineral oil, uniquely barcoded cell specific primers for independent scar and mRNA detection, Spike-in controls and and RNAse inhibitor.
Scartrace
Illumina TruSeq adapaters
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| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina NextSeq 500 |
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| Description |
Scarred GFP
each plate contains 384 sorted cells
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| Data processing |
In scar libraries, first read contains cell barcode (8 first nucleotides) and second read contains the scar. Second reads with a valid cell barcode in first read are mapped to the reference GFP sequence and only reads that map to GFP and contain the reverse PCR primer with at least three mismatches are taken into account for downstream analysis. These reads are mapped again using the pairwise2.align.globalms function from Biopython with match score set to 1; mismatch score to 0.25; open gap penalty to -1; and extending gap penalty to -0.1. Subsequently, we pool scars per cell and by cigar. For each scar library, a double Gaussian is fitted to the distribution of reads per cell to find the threshold in number of reads to select cells. Next, scars were filtered according to their frequency of detection in the full library. For each cell, reads are normalized to 100 and scars representing less than 3.5% are removed. ~
Genome_build: In scar libraries: eGFP sequence extended with ERCC92
Supplementary_files_format_and_content: *scarclones.txt: tabular separated files, with scar percentage (columns) per cell (rows). Cells from the same fish have been named according to barcode ID, plate and organ of origin. Last column (hclust) referes to ID for the clone (cells sharing hclust label have the same scar pattern)
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| Submission date |
Aug 23, 2017 |
| Last update date |
May 15, 2019 |
| Contact name |
Anna Alemany |
| E-mail(s) |
a.alemany@hubrecht.eu
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| Phone |
+31638680750
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| Organization name |
Hubrecht Institue
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| Lab |
AVO
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| Street address |
Uppsalalaan 8
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| City |
Utrecht |
| State/province |
Utrecht |
| ZIP/Postal code |
3584 CT |
| Country |
Netherlands |
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| Platform ID |
GPL20828 |
| Series (1) |
| GSE102990 |
Whole-organism clone-tracing using single-cell sequencing |
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| Relations |
| BioSample |
SAMN07546018 |
| SRA |
SRX3119874 |
| Supplementary data files not provided |
SRA Run Selector |
| Processed data are available on Series record |
| Raw data are available in SRA |
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