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| Status |
Public on Apr 04, 2017 |
| Title |
Male4_Liver |
| Sample type |
SRA |
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| Source name |
male_liver
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| Organism |
Callorhinchus milii |
| Characteristics |
gender: male
developmental stage: Adult
tissue: liver
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| Extracted molecule |
genomic DNA |
| Extraction protocol |
Elephant shark tissue samples were provided by commercial fishermen from recently deceased animals. DNA was purified using a modified magnetic bead approach (Deangelis et al., 1995). Briefly, cells were first homogenised in “GITC” lysis buffer (4 M Guanidine thiocyanate, Sigma G6639; 50 mM Tris, Thermo 15568-025; 20 mM EDTA; Thermo 15575-020; 2% Sarkosyl, Sigma L9150-50G; 0.1% Antifoam, Sigma A8311-50ML) and this lysate mixture was then combined with TE-diluted Sera-Mag Magnetic SpeedBeads (GE Healthcare, GEHE45152105050250) and isopropanol in a volumetric ratio of 2:3:4 respectively. Following capture with a neodymium magnet, beads were washed once with isopropanol, twice with 70% ethanol and resuspended in filter-sterile milliQ H2O.
WGBS-seq was undertaken using a post-bisulfite adapter tagging (PBAT) method adapted from Peat et al., 2014. Briefly, 50-100 ng of purified DNA was subjected to bisulfite conversion using the Imprint DNA modification kit (Sigma, MOD50). Converted DNA underwent first strand synthesis with a biotin labelled adapter sequence possessing seven random nucleotides at its 3’ end (BioP5N7, biotin- ACACTCTTTCCCTACACGACGCTCTTCCGATCTNNNNNNN). The product of first strand synthesis was captured using streptavidin-coated Dynabeads (Thermo, 11205D) and magnetic immobilisation. Double stranded DNA was created using the immobilized first-strand as a template and an additional adapter also possessing seven random nucleotides at its 3’ end (P7N7, GTGACTGGAGTTCAGACGTGTGCTCTTCCGATCTNNNNNNN). Unique molecular barcodes and sequences necessary for binding to Illumina flow-cells were added to libraries by PCR using 1X HiFi HotStart Uracil+ Mix (KAPA, KK2801 and 10 μM indexed Truseq-type oligos, with thermal cycling as follows: 12*(94°C, 80 sec; 65°C, 30 sec; 72°C, 30 sec).
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| Library strategy |
Bisulfite-Seq |
| Library source |
genomic |
| Library selection |
RANDOM |
| Instrument model |
Illumina MiSeq |
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| Data processing |
Trimming was performed to remove poor-quality calls, adapter sequences and 10bp from the 5' end of reads using TrimGalore v0.4.0 (http://www.bioinformatics.babraham.ac.uk/projects/trim_galore/) with default parameters.
Trimmed reads were aligned using Bismark v0.14.3 (Krueger and Andrews, 2011) with the --pbat option specified. Only genome scaffolds of at least 277kb were used for alignment. Bismark mapping reports were used to determine global methylation levels for low coverage elephant shark data.
Genome_build: Callorhinchus milii 6.1.3
Supplementary_files_format_and_content: Bismark mapping report showing mapped calls, with the respective numbers of methylated and unmethylated calls.
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| Submission date |
Apr 03, 2017 |
| Last update date |
May 15, 2019 |
| Contact name |
Tim Hore |
| Organization name |
University of Otago
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| Department |
Department of Anatomy
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| Street address |
270 Great King Street
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| City |
Dunedin |
| ZIP/Postal code |
9016 |
| Country |
New Zealand |
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| Platform ID |
GPL23272 |
| Series (1) |
| GSE96683 |
The methylome of a cartilaginous fish, Callorhinchus milii, reveals conservation of epigenetic regulation across jawed vertebrates |
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| Relations |
| BioSample |
SAMN06678407 |
| SRA |
SRX2702485 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM2562941_Liver_male_4_bismark_SE_report.txt.gz |
780 b |
(ftp)(http) |
TXT |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
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