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Sample GSM1008126 Query DataSets for GSM1008126
Status Public on Aug 26, 2013
Title Mid-maturation-stage embryonic cotyledons
Sample type SRA
Source name Soybean mid- maturation-stage embryonic cotyledons
Organism Glycine max
Characteristics tissue: embyronic cotyledon
developmental stage: mid-maturation
cultivar: Williams 82
Growth protocol Soybean plants (Williams 82) were grown under standard greenhouse conditions (Le et al., PNAS 2010). The mid-maturation (MM) stage seeds are 1.1 - 1.3 cm in length and have a fresh weight between 200 - 250 mg. The tip of the embryonic axis is yellow while the rest of the embryonic axis is green. The embryo was dissected out of the seed coat and the axis was manually separated from the cotyledons of the MM-stage embryo.
Extracted molecule genomic DNA
Extraction protocol Collected tissues were quickly frozen in liquid nitrogen and ground to a fine powder using a mortar and pestle. Genomic DNA was isolated from the powder using the DNEASY Plant Mini kit (Qiagen, Valencia, CA) according to manufacturer’s instructions.Approximately one microgram of genomic DNA was subjected to library preparation following the methods of Hsieh et al. (Hsieh T-F et al. 2009. Science 324:1451-1454) with modifications. We spiked-in ~ three nanogram of unmethylated lambda DNA (Promega) to serve as a control for complete bisulfite conversion. Two libraries were constructed from genomic DNA isolated from seed coat. One library was spiked in ~ three nanogram of unmethylated lambda DNA as all other libraries; the other library was spiked in ~ three nanogram of methylated lambda DNA to serve as a control for over conversion (i.e. conversion of methylated cytosine to uracil) during the bisulfite conversion step. The methylated lambda DNA was generated by treating unmethylated lambda DNA with CpG Methyltransferase (New England BioLabs). Adapter-ligated genomic DNA was subjected to two rounds of bisulfite (BS) treatment using the EpiTect kit (Qiagen, Valencia, CA). BS-treated DNA was purified using AMpure XP beads (Beckman) and PCR-amplified for 10 cycles using ExTaq (EpiCentre) DNA polymerase. PCR-amplified DNA fragments were size selected using the AMpure XP beads (Beckman). Phi-X174 DNA was spiked in to the library by the sequencing facility before cluster formation and sequencing.
Library strategy Bisulfite-Seq
Library source genomic
Library selection RANDOM
Instrument model Illumina HiSeq 2000
Description Whole embryonic cotyledons were collected from mid-maturation stage embryos. Seeds containing mid-maturation embryos are 1.1 - 1.3 cm in length and have a fresh weight between 200 - 250 mg.
Data processing Basecalls performed using RTA version
Original data file from the Illumina sequencing pipeline. Each lane of sequencing is attached as an individual compressed file. The sequence data are in the QSEQ format
We aligned the raw reads to a pre-processed reference genome using BS Seeker [Chen et al. BMC Bioinformatics (2010)] allowing for two mismatches. The pre-processed reference genome consisted of sequences from the soybean genome (Glyma version 1.0.1) [Schmutz et al. Nature (2010)] obtained from the Phytozome website (, soybean chloroplast genome (GenBank: DQ317523), lambda reference genome (GenBank: J02459), and Phi-X174 reference genome (GenBank: J02482).
Reads containing three consecutive methylation in the non-CG sites were removed, possibly representing non-converted cytosines (Cokus et al. Nature 2008). Clonal reads possibly arising during the PCR amplification step were collapsed into one read.
Methylation level of sampled cytosine was calculated as (methylated calls / (methylated calls + unmethylated calls)).
Genome_build: Glyma version 1.01
Supplementary_files_format_and_content: tab-delimited text file including methylation level of each sampled cytosine
Submission date Sep 21, 2012
Last update date Aug 26, 2013
Contact name Bob Goldberg
Phone 310-825-3270
Organization name University of California, Los Angeles
Department Molecular, Cell and Developmental Biology
Street address 610 Charles E Young Drive East
City Los Angeles
State/province CA
ZIP/Postal code 90095
Country USA
Platform ID GPL15008
Series (1)
GSE41061 Methylation Changes in Soybean Mid-Maturation Seed Parts
SRA SRX189100
BioSample SAMN01180425

Supplementary file Size Download File type/resource
GSM1008126_wm.b1.cot.bsseq.chr1-20.perC.txt.gz 1.0 Gb (ftp)(http) TXT
Raw data provided as supplementary file
Processed data provided as supplementary file

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