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Status |
Public on Dec 05, 2019 |
Title |
MeDIP-seq of DNA from pig-308 |
Sample type |
SRA |
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Source name |
Adipocytes
|
Organism |
Sus scrofa |
Characteristics |
cell type: adipocyte Sex: M phenotype: obese
|
Treatment protocol |
Mature adipocytes were isolated from approx. 15 g of the retroperitoneal adipose tissue from each animal, treated with 0.2% collagenase and filtered through a 200-um nylon filter.
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Extracted molecule |
genomic DNA |
Extraction protocol |
DNA was extracted from the mature adipocytes using the Epicenter kit (Illumina), and total RNA was isolated from the mature adipocytes according to the protocol described in (Cirera, 2013) using TriReagent and included a DNAse treatment step. RNA libraries were prepared for sequencing using Truseq lib prep kit from Illumina together with RiboZero Gold using 0.5ug of total RNA as input. mRRBS libraries were prepared as described in Boyle (2012) using 200ng DNA as input. MeDiP libraries were prepared from 1.1ug DNA, fragmented in a bioruptor and methylated DNA fragments were enriched using MethylMiner Methylated DNA Enrichment KIt (cat#ME10025). Sequencing libraries were generated according to the low Input TruSeq library preparation protocol.
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Library strategy |
MeDIP-Seq |
Library source |
genomic |
Library selection |
5-methylcytidine antibody |
Instrument model |
Illumina HiSeq 2000 |
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Description |
data_all.gt.Obese_lean.diffReps.tsv data_all.gt.Obese_lean.diffReps.hotspot.tsv meDIP-308
|
Data processing |
RRBS: The raw reads were cleaned for quality and adapters using and in-house tools removing TrueSeq adapters and clipping read tails of phred quality less than 20. The reads were mapped to the pig genome version 10.2 using bismark v 0.10 (Krueger and Andrews, 2011) with standard parameters except using bowtie 2 v2.1.0 for the mapping option of bismark (Langmead and Salzberg, 2012). The mapped reads were analyzed using Metilene v 0.2-7 requiring at least five pigs in both the obese and lean groups (–X 5 –Y 5) mDNAcap: The raw reads were trimmed for quality and adapters using trimmomatic V0.22 (Bolger et al., 2014) (quality trimming by phred quality of 20) The cleaned reads were mapped to the pig genome version 10.2 using segemehl V0.1.4 (Hoffmann et al., 2009) and the following parameters: accuracy 95%, and minimum mapping size 18. The mapped reads were analyzed with diffReps V1.55.4 using the G-test and default values for all remaining parameters RNAseq: The raw reads were cleaned for quality and adapters using and in-house tools removing TrueSeq adapters and clipping read tails of phred quality less than 20 The reads were mapped to the pig transcriptome using the STAR aligner v2.3.1 (Dobin et al., 2013) with the following options and data: pig genome version 10.2, ensembl annotation version 70 and with intron-motifs [XS:A] tags in the SAM output. The aligned reads were assigned to transcripts of the ensembl version 70 annotation with cufflinks version 2.2.0 (Trapnell et al., 2013). The subsequent differential expression analysis was performed with cuffdiff from the cufflinks package and was based on the Ensembl v70 annotation. Standard options were used for the cufflinks programs in all cases. All further analysis was done using the annotation from ensemble v81. Supplementary_files_format_and_content: [RNAseq] fpkm_tracking.tsv, gene_exp.diff.tsv (1=obese, q2=lean) Supplementary_files_format_and_content: [MeDIP-seq] data_all.gt.Obese_lean.*.tsv (Treatment=obese, control=lean) Supplementary_files_format_and_content: [RRBS-seq] metilene.*.tsv
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Submission date |
Sep 03, 2019 |
Last update date |
Dec 05, 2019 |
Contact name |
Christian Anthon |
E-mail(s) |
anthon@rth.dk
|
Phone |
+4521510929
|
Organization name |
University of Copenhagen
|
Department |
Veterinary and Animal Sciences
|
Lab |
RTH
|
Street address |
Thorvaldsens Vej 57
|
City |
Frederiksberg C |
ZIP/Postal code |
1871 |
Country |
Denmark |
|
|
Platform ID |
GPL11429 |
Series (1) |
GSE136754 |
Epigenetic and transcriptomic characterization of pure adipocyte fractions from obese pigs identifies candidate pathways controlling metabolism |
|
Relations |
BioSample |
SAMN12683973 |
SRA |
SRX6793315 |