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| Status |
Public on Nov 01, 2020 |
| Title |
healthy2_ip |
| Sample type |
SRA |
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| Source name |
human heart tissue
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| Organism |
Homo sapiens |
| Characteristics |
disease state: healthy heart
tissue: heart
isolation method: m6A-IP RNA
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| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was purified with TRIzol reagents from human heart samples. Polyadenylated RNA was then isolated from total RNA with two rounds of polyA tail purification using Dynabeads® mRNA DIRECT™ kit (Thermo Scientific). rRNA was further removed from polyadenylated RNA using RiboMinus Eukaryote kit (Thermo Scientific). 50 ng polyadenylated RNA after rRNA depletion was digested by nuclease P1 (Sigma, N8630), respectively, in 20 μl of buffer containing 25 mM NaCl and 2.5 mM ZnCl2 for 1 h at 42 °C. 1 ug polyadenylated RNA was adjusted to 10 ng/μl in 100 μl and fragmented using a BioRuptor ultrasonicator (Diagenode) with 30 s on/off for 30 cycles. After fragmentation, 5% of the fragmented RNA was saved as input. m6A marked fragments were enriched using EpiMark N6-Methyladenosine Enrichment Kit (NEB) following the manufacturer’s protocols and collected as IP. Both input and IP samples were subjected to library preparation using TruSeq Stranded mRNA Library Prep Kit (Illumina) following the manufacturer’s protocols
Both input and IP samples were subjected to library preparation using TruSeq Stranded mRNA Library Prep Kit (Illumina) following the manufacturer’s protocols.
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| Library strategy |
RIP-Seq |
| Library source |
transcriptomic |
| Library selection |
other |
| Instrument model |
Illumina HiSeq 2000 |
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| Data processing |
alignment of fastq files using hisat2 v2.2.0 with default settings
count matricies formed using htseq v2.0 with default settings
input differential expression calculation using DESeq2, FC set to 1.5 and p-val set to 0.05.
m6a peak calling using exomePeak v1.6, FDR set to 0.05
Genome_build: hg19
Supplementary_files_format_and_content: hisat2 alignment file (.bam), htseq count matrix (.txt), DEseq2 analysis (.csv), m6A exomePeak (.bed)
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| Submission date |
Oct 08, 2020 |
| Last update date |
Nov 02, 2020 |
| Contact name |
Federica Accornero |
| E-mail(s) |
federica.accornero@osumc.edu
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| Organization name |
Ohio State University Medical Center
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| Street address |
473 W 12th Ave.
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| City |
Columbus |
| State/province |
OH |
| ZIP/Postal code |
43210 |
| Country |
USA |
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| Platform ID |
GPL11154 |
| Series (1) |
| GSE159243 |
Remodeling of the m6A landscape in the heart reveals conserved post-transcriptional events underlying cardiomyocyte hypertrophy |
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| Relations |
| BioSample |
SAMN16397814 |
| SRA |
SRX9264962 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM4824283_healthy2_IP.txt.gz |
236.0 Kb |
(ftp)(http) |
TXT |
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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