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Sample GSM2182806 Query DataSets for GSM2182806
Status Public on Nov 30, 2017
Title Aedes_aegypti_RE
Sample type SRA
 
Source name whole mosquitoes
Organism Aedes aegypti
Characteristics treatment: Wild type
strain: red eye strain
Extracted molecule genomic DNA
Extraction protocol Genomic DNA was prepared using the DNeasy Blood & Tissue Kit (Qiagen) following the manufacturer's instructions
5 µg (8 % spike-in control DNA of human blood & 1 % spike-in control unmethylated bacteriophage lambda DNA) of high molecular weight DNA were used for fragmentation using the Covaris S2 AFA System in a total volume of 100 µl. Fragmentation-run parameters: Duty cycle 10%; Intensity: 5; Cycles/burst: 200; Time: 3 min; number of cycles: 3, resulting in a total fragmentation-time of 180 s. Fragmentation was confirmed with a 2100 Bioanalyzer (Agilent Technologies) using a DNA1000 chip. The fragmented DNAs were concentrated to a final volume of 75 µl using a DNA Speed Vac. End repair of fragmented DNA was carried out in a total volume of 100 µl using the Paired End DNA Sample Prep Kit (Illumina) as recommended by the manufacturer. For the ligation of the adaptors, the Illumina Early Access Methylation Adaptor Oligo Kit and the Paired End DNA Sample Prep Kit (Illumina) were used, as recommended by the manufacturer. For the size selection of the adaptor-ligated fragments, we used the E-Gel Electrophoresis System (Invitrogen) and a Size Select 2% precast agarose gel (Invitrogen). Each fragmented DNA was loaded on two lanes of the E-gel. Electrophoresis was carried out using the “Size Select” program for 16 min. According to the standard loaded (50 bp DNA Ladder, Invitrogen), 240 bp fragments were extracted from the gel, pooled, and directly transferred to bisulfite treatment without further purification. For the bisulfite treatment we used the EZ-DNA Methylation Kit (Zymo) as recommended by the manufacturer with the exception of a modified thermal profile for the bisulfite conversion reaction. The conversion was carried out in a thermal cycler using the following thermal profile: 95°C for 15 s, 50°C for 1 h, repeat from step 1, 15×, 4°C for at least 10 min. The libraries were subsequently amplified, using the Fast Start High Fidelity PCR System (Roche) with buffer 2, and Illuminas PE1.1 and PE2.1 amplification primers. PCR thermal profile: 95°C for 2 min, 95°C for 30 s, 65°C for 20 s, 72°C for 30 s, then repeat from step 2, 11×, 72°C for 7min, hold at 4°C. PCR reactions were purified on PCR purification columns (MinElute, Qiagen) and eluted in 20 µl elution buffer (Qiagen).
 
Library strategy Bisulfite-Seq
Library source genomic
Library selection RANDOM
Instrument model HiSeq X Five
 
Data processing Illumina Casava1.8.1 software used for basecalling.
Preprocessing, read trimming: shore 0.6.2
Preprocessing, quality filtering: shore 0.6.2
BS-mapping: bsmap 2.0
computation of methylation ratios: methratio.py (part of bsmap package)
Genome_build: AaegL3
Supplementary_files_format_and_content: bedGraph-format containing the genomic methylation ratios and coverage of the sample
 
Submission date May 31, 2016
Last update date May 15, 2019
Contact name Cassandra Falckenhayn
E-mail(s) c.falckenhayn@dkfz-heidelberg.de
Organization name German Cancer Research Center (DKFZ)
Department Epigenetics
Street address Im Neuenheimer Feld 580
City Heidelberg
ZIP/Postal code 69120
Country Germany
 
Platform ID GPL21967
Series (1)
GSE82063 Comprehensive DNA methylation analysis of the Aedes aegypti genome
Relations
BioSample SAMN05190535
SRA SRX1810236

Supplementary file Size Download File type/resource
GSM2182806_Aaedes_RE_cov.bedgraph.gz 542.8 Mb (ftp)(http) BEDGRAPH
GSM2182806_Aaedes_RE_ratio.bedgraph.gz 485.4 Mb (ftp)(http) BEDGRAPH
SRA Run SelectorHelp
Raw data are available in SRA
Processed data provided as supplementary file

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