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Sample GSM2124006

Query DataSets for GSM2124006

Status

Public on Jun 24, 2016

Title

Mock_treated_4

Sample type

SRA

Source name

Mock treated 4

Organism

Arabidopsis thaliana

Characteristics

ecotype: coumbia-11
tissue: whole seedling
treatment: 0 uM

Growth protocol

Agarose gel (Ameresco) with added half-strength Linsmaier and Skoog nutrients (Caisson Laboratories, Inc) was prepared and autoclaved. 5-azacytidine (Sigma) and zebularine (APExBIO) were dissolved in dimethyl-sulfoxide (DMSO) and water, respectively, before being added to the liquefied cooling agar at final concentrations of 25 µM, 50 µM and 100 µM. Columbia-0 (col-0) background Arabidopsis thaliana seeds were then subjected to a ethanol-based seed sterilization and approximately 30 seeds were plated on the solid agar. As a control, seeds were plated on agar containing DMSO with no chemical demethylating agent (AZA mock-treated control) and agar containing no DMSO or chemical demethylating agent (untreated control). Three replicates of each treatment type were simultaneously plated and grown in parallel. After a two-day stratification period at 4ºC, the seeds were removed to room temperature and allowed to grow for 8 days under constant light.

Extracted molecule

total RNA

Extraction protocol

Tissue was flash frozen in liquid nitrogen for all experiments including RNA-seq and MethylC-seq. DNA was isolated using a Qiagen Plant DNeasy kit (Qiagen, Valencia, CA) following the manufacturer’s recommendations. RNA was isolated using TRIzol (Thermo Scientific, Waltham, MA) following the manufacturer’s instructions.
RNA-seq: RNA-seq libraries were constructed using Illumina TruSeq Stranded RNA LT Kit (Illumina, San Diego, CA) following the manufacturer’s instructions with limited modifications. The starting quantity of total RNA was adjusted to 1.3 µg, and all volumes were reduced to a third of the described quantity.
MethylC-seq libraries were constructed using the MethylC-seq protocol (Urich et al. 2015).

Library strategy

RNA-Seq

Library source

transcriptomic

Library selection

cDNA

Instrument model

Illumina NextSeq 500

Description

RNA Sequencing on Mock treated 4

Data processing

Data were mapped to the TAIR 10
MethylC-seq analysis were performed as described in Schmtiz et al. Epigenome-wide inheritance of DNA methylation variants in a recombinant inbred population. Genome Research (2013).
Genome_build: TAIR 10
Supplementary_files_format_and_content: MethylC-seq data (marked as “allc_sampel.tsv”) column 1 = chr = chromosome; column 2 = pos = coordinate for the cytosine position on the chromosome; column 3 = strand = + or - strand; column 4 = mc_class = context of the cytosine and the two following bases from the same strand; column 5 = mc_count = number of reads supporting a methylated cytosine; column 6 = total = total number of reads at that position; column 7 = methylated = cytosine is considered methylated if there is a 1 or unmethylated if there is a 0
Supplementary_files_format_and_content: RNA-seq data (marked as "gene_exp.diff") column 1 = Tested id = A unique identifier describing the transcipt, gene, primary transcript, or CDS being tested; column 2 = gene = The gene_name(s) or gene_id(s) being tested; column 3 = locus = Genomic coordinates for easy browsing to the genes or transcripts being tested.; column 4 = sample 1 = Label (or number if no labels provided) of the first sample being tested; column 5 = sample 2 = Label (or number if no labels provided) of the second sample being tested; column 6 = Test status = Can be one of OK (test successful), NOTEST (not enough alignments for testing), LOWDATA (too complex or shallowly sequenced), HIDATA (too many fragments in locus), or FAIL, when an ill-conditioned covariance matrix or other numerical exception prevents testing.; column 7 = FPKMx = FPKM of the gene in sample x; column 8 = FPKMy = FPKM of the gene in sample y; column 9 = log2(FPKMy/FPKMx) = The (base 2) log of the fold change y/x; column 10 = test stat = The value of the test statistic used to compute significance of the observed change in FPKM; column 11 = p = The uncorrected p-value of the test statistic; column 12 = q = The FDR-adjusted p-value of the test statistic; column 13 = significant = Can be either “yes” or “no”, depending on whether p is greater then the FDR after Benjamini-Hochberg correction for multiple-testing

Submission date

Apr 14, 2016

Last update date

May 15, 2019

Contact name

Robert J Schmitz

E-mail(s)

schmitz@uga.edu

Organization name

University of Georgia

Department

Genetics