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| Status |
Public on Oct 19, 2023 |
| Title |
L571-5hmU_chemical_mapping_harsh_oxid |
| Sample type |
SRA |
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| Source name |
L571-5hmU_chemical_mapping_harsh_oxid
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| Organism |
Breviolum minutum |
| Characteristics |
tissue: whole dinoflagellate
sample type: B. minutum 5-hmU chemical mapping
chip antibody: n/a
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| Growth protocol |
(B. minutum) The clonal axenic Symbiodinium/Breviolum minutum strain SSB01 was used. Stock cultures were grown in Daigo's IMK medium for marine microalgae (Wako Pure Chemicals) supplemented with casein hydrolysate (IMK+Cas) at 27C at a light intensity of 10 µmol photons m-2s-1 from Philips ALTO II 25-W bulbs on a 12-h-light:12-h-dark cycle. The medium was prepared in artificial seawater (ASW).
(yeast) Standard yeast cell culture practices were followed. Cells were grown in YPD media at 30C until harvesting.
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| Extracted molecule |
genomic DNA |
| Extraction protocol |
Chemical mapping of 5-hmU as carried out following the previously described by Kawasaki et al. chemical conversion method. Briefly, sheared genomic DNA was used as input and end prep and adapter ligation were carried out using the NEBNext Ultra II DNA Library Prep Kit. After the ligation step, DNA was purified using AMPure XP beads and eluted in 50 µL of H2O. DNA denaturation was performed by adding NaOH to a final concentration of 0.05M and incubating at 37◦C for 30 minutes. Oxidation was carried out by adding 2 µL of KRuO4 solution (15 mM in 0.05 M NaOH) and incubating for 30 minutes at room temperature. Oxidized DNA was purified using AMPure XP beads and extension was carried out by mixing 13.5 µL DNA, 1.6 µL 100 mM MgSO4, 2 µL NEB Index Primer, 2 µL 10× ThermoPol Reaction Buffer (NEB), 0.5 µL 10 mM dNTP mix, and 0.4 µL Bst DNA Polymerase, Large Fragment (NEB), then incubating for 1 hour at 37◦C
PCR amplification was carried out using the NEB Ultra DNA Library Prep Kit, with 12 cycles of PCR. Final libraries were purified using AMPure XP beads.
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| Library strategy |
OTHER |
| Library source |
genomic |
| Library selection |
other |
| Instrument model |
NextSeq 550 |
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| Description |
L571-5hmU_chemical_mapping_harsh_oxid
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| Data processing |
(read alignment, ATAC-seq and MeDIP-seq data) Bowtie 1.0.1
(bigWig file generation) custom code
(read alignment, SMF data) bwa-meth
(read alignment, 5-hmU chemical conversion data) slamdunk
Assembly: custom and GCA_000507305.1_ASM50730v1 for Breviolum minutum, hg38 for human, sacCer3 for S. cerevisiae; C_glabrata_CBS138 for Candida albicans
Supplementary files format and content: bedGraph of read coverage, normalized to RPM for bigWig files for ATAC-seq and MeDIP-seq; slamdunk output for 5-hmU conversion datasets; MethylDacket bedGraph output for SMF datasets
Library strategy: 5-hmU chemical mapping
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| Submission date |
Aug 30, 2023 |
| Last update date |
Oct 19, 2023 |
| Contact name |
Georgi Kolev Marinov |
| Organization name |
STANFORD UNIVERSITY
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| Department |
Genetics
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| Street address |
279 Campus Drive West, Beckman Center, B-257A/259
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| City |
Stanford |
| State/province |
California |
| ZIP/Postal code |
94305-5101 |
| Country |
USA |
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| Platform ID |
GPL33721 |
| Series (1) |
| GSE241969 |
Genome-wide distribution of 5-hydroxymethyluracil and chromatin accessibility in the Breviolum minutum genome |
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| Relations |
| BioSample |
SAMN37205250 |
| SRA |
SRX21542009 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM7746910_L571-5hmU_harsh_oxid.trimmed.end1+end2.fastq_slamdunk_mapped_filtered_tcount_plus.bigWig |
465.4 Mb |
(ftp)(http) |
BIGWIG |
SRA Run Selector |
| Raw data are available in SRA |
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