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| Status |
Public on May 07, 2015 |
| Title |
Organoid 2 derived from patient 20 |
| Sample type |
SRA |
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| Source name |
Colonic Adenocarcinoma derived organoid
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| Organism |
Homo sapiens |
| Characteristics |
sample type: Organoid culture
tissue: Colonic Adenocarcinoma derived organoid
protocol: CEL-seq (Hashimshony et al., Cell reports 2012)
subject: patient 20
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| Growth protocol |
Organoids derived from colorectal cancer cases were resuspended in matrigel. Subseqeuntly, droplets of matrigel (10 ul), were put in a dish. After letting the Matrigel solidy, Human Intestinal Stem Cell medium (HISC) (Basal culture medium with 50% Wnt conditioned medium, 20% R-Spondin conditioned medium, 10% Noggin conditioned medium, 1 x B27, 1,25 mM n-Acetyl Cysteine, 10 mM Nicotinamide, 50 ng/ml human EGF, 10 nM Gastrin, 500 nM A83-01, 3 uM SB202190, 10 nM Prostaglandine E2 and 1 x Primocin (Vivogen)) minus Wnt was added and the cells were incubated at 37 oC.
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| Extracted molecule |
total RNA |
| Extraction protocol |
For single organoid mRNA sequencing, organoids were hand-picked and deposited into trizol (life technologies).
The RNA was extracted using chloroform and precipitated with iso-propanol. The RNA-pellet was then processed using the CEL-seq protocol (Hashimshony et al., 2012) and sequenced on an Illumina Nextseq using 75bp paired end sequencing.
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| Library strategy |
OTHER |
| Library source |
transcriptomic |
| Library selection |
other |
| Instrument model |
Illumina NextSeq 500 |
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| Description |
mRNA
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| Data processing |
After sequencing, read 1 was aligned to the hg19 RefSeq human transcriptome downloaded from the UCSC genome browser (Meyer et al., 2013) using bwa (Li and Durbin, 2010) with default parameters. Read 2 contains a barcode identifying the sample from which the read originated. CEL-seq only sequences the most 3’ prime end of a transcript and generates one read per transcript. Samples with less than 500.000 reads were discarded after which the remaining samples were rpm normalized.
Genome_build: hg19
Supplementary_files_format_and_content: mRNA count data for all samples, each column is a sample each row is a gene
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| Submission date |
Jan 23, 2015 |
| Last update date |
May 15, 2019 |
| Contact name |
Lennart Kester |
| E-mail(s) |
l.kester@hubrecht.eu
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| Organization name |
Hubrecht Institute
|
| Street address |
Uppsalalaan 8
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| City |
Utrecht |
| ZIP/Postal code |
3584CT |
| Country |
Netherlands |
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| Platform ID |
GPL18573 |
| Series (1) |
| GSE65253 |
Prospective derivation of a 'Living Organoid Biobank' of colorectal cancer patients |
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| Relations |
| BioSample |
SAMN03292596 |
| SRA |
SRX852083 |
| 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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