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Sample GSM2332658

Query DataSets for GSM2332658

Status

Public on Nov 22, 2016

Title

46_BS-seq_needle_rep1

Sample type

SRA

Source name

needle

Organism

Picea abies

Characteristics

clone: Z4006
cell type: needle
Stage: young needle

Extracted molecule

genomic DNA

Extraction protocol

DNA was extracted from needles of the sequenced Z4006 Picea abies genotype, as described in Nystedt et al 2013 Nature. For the SE culture sample, material was collected at the proliferation stage, as described in Businge et al 2012 Tree Physiol, and DNA extracted as for the needle samples. The culture was generated from seeds obtained from the sequenced ‘Z4006’ genotype.
BS-seq libraries were prepared using the TruSeq DNA LT kit (Illumina) as described previously Du J et al., (2014) Mol Cell, except that EZ DNA Methylation-Lightning Kit (Qiagen) was used for bisulfite conversion of the DNA. BS-Seq libraries were sequenced on a HiSeq 2500 system (Illumina) to obtain single-end 100bp reads per manufacture instructions.
For BSPCR-seq, genomic DNA were bisulfite treated by EZ DNA Methylation-Lightning Kit (Zymo) and then amplified by locus-specific primers. Amplification products were used for library generation. Libraries in repliate 1 were generated by Ovation Ultralow V2 kit (NuGen) and libraries in replicate 2 were generated by TruSeq Nano DNA LT kit (illumina). BSPCR-seq libraries were sequenced on a HiSeq 2000 system (Illumina) to obtain single-end 100bp reads per manufacture instructions.

Library strategy

Bisulfite-Seq

Library source

genomic

Library selection

RANDOM

Instrument model

Illumina HiSeq 2500

Data processing

BS-seq libraries were sequenced on a HiSeq 2500 system (Illumina) to obtain single-end 100 bp reads.
All BS-seq reads were aligned against the Norway spruce reference genome, as well as the chloroplast genome, using Bismark v0.13.0 with following parameters: -q --score_min L,0,-0.3 -most_valid_alignments 1 --bowtie2.
For BSPCR-seq analysis, only reads with correct BSPCR primers were kept. Primers were then trimmed with custome perl script. Then reads were alinged to the Norway spruce reference genome, as well as the chloroplast genome, using BSMAP(version 2.74) by allowing 2 mismatches(-v2) unique mapped reads (-w1). Then only cytosines over correct PCR regions and PCR strands were kept. Methylation ratio were calculated using #C/(#C+#T).

Genome_build: Norway spruce reference genome.
Supplementary_files_format_and_content: wig files were generated by custom perl script, and show each cytosine methylation levels by #C/(#C+#T).

Submission date

Sep 29, 2016

Last update date

May 15, 2019

Contact name

Haifeng Wang

E-mail(s)

[email protected]

Organization name

Fujian Agriculture and Forestry University

Street address

Shangxia dian