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Sample GSM2306322 Query DataSets for GSM2306322
Status Public on Dec 12, 2017
Title WT_meiocyte_BSseq_Rep2
Sample type SRA
 
Source name WT_meiocyte_BSseq
Organism Arabidopsis thaliana
Characteristics ecotype background: Col-0
genotype/variation: wild type
age: 5-6 week
tissue/cell type: male meiocyte
Growth protocol A. thaliana plants of Col-0 ecotype were grown under 16h light/ 8h dark in a growth chamber.
Extracted molecule genomic DNA
Extraction protocol Stage 9 flower buds were collected and gently squeezed between a glass slide and coverslip. The released meiocytes (of meiotic prophase I) were examined carefully under a microscope. Clean meiocytes free from any somatic cell debris were transferred to a new slide using capillary glass pipettes, washed by 1 x PBS buffer 3 times, and frozen in liquid nitrogen. Sperm cells and vegetative cell nuclei were isolated by Fluorescence-activated cell sorting (FACS)
Single-end bisulfite sequencing libraries for Illumina sequencing were constructed using the Ovation Ultralow Methyl-Seq Library Systems (Nugen, #0336) and EpiTect Fast Bisulfite Conversion (Qiagen, #59802) kits according to the kit protocols, except the incorporation of two rounds of bisulfite conversion.Strand-specific RNA sequencing libraries were prepared using ScriptSeq v2 RNA-Seq (Illumina, #SSV21106) Library Preparation and Ovation RNA-Seq Systems for Model Organisms Arabidopsis (Nugen, #0351) kits
 
Library strategy Bisulfite-Seq
Library source genomic
Library selection RANDOM
Instrument model Illumina NextSeq 500
 
Data processing Bisulfite-Seq: We used Perl scripts to convert all the Cs in the reads (and in the scaffolds) to Ts, and aligned the converted reads to the converted reference scaffold, allowing up to two mismatches per read. 76 base reads were divided into the first 45 and the last 31 bases, 100 base reads and 150 base reads were divided in half. Each half of the read was aligned independently using bowtie, allowing up to two mismatches. The coordinates of the two halves were subsequently correlated; the second half was discarded if it did not match the first.
Single_C: We used Perl scripts to recover the original sequence of each mapped read and, for each C (on either strand), count the number of times it was sequenced as a C or a T.
50bp_window: We used a Perl script to calculate fractional methylation (#C/(#C+#T)) within a 50 bp sliding window for each sequence context (CG, CHG, CHH).
RNASeq: Tophat-2.0.10 followed by Cufflinks-2.2.1 were used to highlight differentially expressed genes between samples.
Genome_build: TAIR10
Supplementary_files_format_and_content: All files are in GFF format. Files contain fractional methylation data either for individual cytosines (single-c) or in 50 bp windows.
 
Submission date Sep 08, 2016
Last update date May 15, 2019
Contact name Xiaoqi Feng
E-mail(s) xiaoqi.feng@jic.ac.uk
Organization name John Innes Centre
Department Cell and Developmental Biology
Lab Xiaoqi Feng
Street address Norwich Research Park
City Norwich
State/province Norfolk
ZIP/Postal code NR4 7UH
Country United Kingdom
 
Platform ID GPL19580
Series (1)
GSE86583 Sexual-lineage-specific DNA methylation regulates meiosis in Arabidopsis
Relations
BioSample SAMN05751924
SRA SRX2148721

Supplementary file Size Download File type/resource
GSM2306322_WT_meiocyte_rep2.CG.tair10.gff.gz 34.9 Mb (ftp)(http) GFF
GSM2306322_WT_meiocyte_rep2.CG.tair10.w50.gff.gz 17.5 Mb (ftp)(http) GFF
GSM2306322_WT_meiocyte_rep2.CHG.tair10.gff.gz 38.8 Mb (ftp)(http) GFF
GSM2306322_WT_meiocyte_rep2.CHG.tair10.w50.gff.gz 17.8 Mb (ftp)(http) GFF
GSM2306322_WT_meiocyte_rep2.CHH.tair10.gff.gz 172.3 Mb (ftp)(http) GFF
GSM2306322_WT_meiocyte_rep2.CHH.tair10.w50.gff.gz 21.2 Mb (ftp)(http) GFF
SRA Run SelectorHelp
Raw data are available in SRA
Processed data provided as supplementary file

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