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| Status |
Public on Aug 24, 2020 |
| Title |
scRCAT-seq_single_hESC_13 |
| Sample type |
SRA |
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| Source name |
hESC
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| Organism |
Homo sapiens |
| Characteristics |
strain: H9
tissue: hESC
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| Extracted molecule |
polyA RNA |
| Extraction protocol |
DRG, oocytes, hESC, HEK293T, ARPE, mESC, hESC_drived_organoid were dissected and dissociated into single cells. hESC, HEK293T, ARPE, mESC, hESC_drived_organoid were dissected respectively and mixed together.
The scRCAT-seq library was prepared according to the scRCAT-seq protocol
The smart-seq2 library was prepared according to the smart-seq2 protocol
The Iso-Seq library was prepared according to the Isoform Sequencing protocol
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| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
HiSeq X Ten |
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| Description |
applying single cell method to hESC
hESCsccatUMI_5.bed.gz, hESCsccatUMI_3.bed.gz
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| Data processing |
Illumina Casava1.7 software used for basecalling.
5' and 3' sequences were extracted from paired-end sequences using Perl, Python and shell script
5' and 3' sequences were aligned to mm10, using STAR (2.6.1a), with with parameters (--outFilterMultimapNmax 1 --outFilterScoreMinOverLread 0.6 --outFilterMatchNminOverLread 0.6).
Peaks were called with CAGEr (v 1.24.0) .
For smart-seq2 data, sequenced reads were trimmed for adaptor sequence using cutapapt (v 1.18) and aligned to mm10 using STAR with with parameters as described above.
For Iso-seq data, sequence data were processed using the SMRTlink5.0 software. Circular consensus sequence (CCS) was generated from subread BAM files, parameters: min_length 200, max_drop_fraction 0.8, no_polish TRUE, min_zscore -9999, min_passes 1, min_predicted_accuracy 0.8, max_length 18000. CCS.BAM files were classified into full length and non-full length reads using pbclassify.py script, ignore polyA false, minSeq Length 200. Non-full length and full-length fasta files produced were then fed into the cluster step, which does isoform-level clustering(ICE), followed by final Arrow polishing, hq_quiver_min_accuracy 0.99, bin_by_primer false, bin_size_kb 1, qv_trim_5p 100, qv_trim_3p 30.
Additional nucleotide errors in consensus reads were corrected using the Illumina RNA seq data with the software LoRDEC.
consensus reads were aligned to reference Genome using GMAP with parameters --no-chimeras --cross-species --expand-offsets 1 –B 5 –K 50000 –f samse –n 1 against reference genome.
Gene structure analysis was performed using TAPIS pipeline.
Genome_build: hg38 and mm10
Supplementary_files_format_and_content: [scRCAT-seq] *bed file reports the TSS/TES position
Supplementary_files_format_and_content: [Iso-seq] *gtf reports the isoform information which were detected by Iso-seq.
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| Submission date |
Jun 01, 2020 |
| Last update date |
Aug 29, 2020 |
| Contact name |
Jiawei Zhong |
| Organization name |
Karolinska Institutet
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| Department |
Department of Medicine, Huddinge
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| Lab |
Mikael Rydén & Niklas Mejhert lab
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| Street address |
Blickagången 16, Flemingsberg
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| City |
Stockholm |
| ZIP/Postal code |
14182 |
| Country |
Sweden |
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| Platform ID |
GPL20795 |
| Series (1) |
| GSE134311 |
scRCAT-seq: simultaneously profile RNA TSS and TES at single-cell level |
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| Relations |
| BioSample |
SAMN15077061 |
| SRA |
SRX8450069 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM4586169_hESC_8N_H15_3.bed.gz |
576.2 Kb |
(ftp)(http) |
BED |
| GSM4586169_hESC_8N_H15_5.bed.gz |
148.4 Kb |
(ftp)(http) |
BED |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
| Processed data are available on Series record |
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