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Sample GSM7765162

Query DataSets for GSM7765162

Status

Public on Sep 16, 2023

Title

GmMM-Input-br2

Sample type

SRA

Source name

embryos

Organism

Glycine max

Characteristics

tissue: embryos
genotype: Williams 82
cell type: mid-maturation
chip antibody: none

Treatment protocol

Mid-maturation stage embryos were dissected from seeds weighing between 200 and 250 mg. Tissues were crosslinked under vaccuum for 10 minutes in Buffer A (0.4M sucrose, 10 mM Tris pH 8.0, 1mM EDTA pH 8.0) containing 1% formaldehyde, at room temperature, then quenched in 100 mM Glycine for 5 minutes. Tissues were rinsed twice with deionized water and stored at -80C until use.

Growth protocol

Soybean cv Williams 82 plants were grown in 2 gallon pots containing Ron’s mix (1 part coarse sand, 1 part compost, 1 part peat moss, 3 pounds/yard Dolomite) in UC Davis greenhouses at temperatures ranging 22-30°C and under 16 hour-days.

Extracted molecule

genomic DNA

Extraction protocol

Chromatin isolation and immunoprecipitation were performed as described in Gendrel AV, Lippman Z, Martienssen R, Colot V. Nat Methods. 2005;2:213–218, except that the isolated nuclei were resuspended in lysis buffer described in by Johnson et al. (2002) and sonicated to achieve chromatin fragments ranging from 100 to 500 bp. Nuclear extracts were either set aside for whole genome extracts (input control) or incubated overnight with 2 ug of an antibody raised against the peptide CTEKGVIRKEPSLPRQ, or 3 ug of an antibody raised against CSGYAGNGKRDVGYPP, corresponding to the amino acids 103-117 and 275-289 of GLYMA.13G153200, respectively. Crosslinks were reversed and protein digested as described in Dahl and Collas, Nat Protoc. 2008;3(6):1032-45, extracted with phenol:chloroform and precipitated using standard procedures.
ChIP and Input libraries were generated using Nugen Ovation Ultralow System V2, following the manufaturer's instructions and enriched by PCR using 15 cycles. Libraries were size-selected by electrophoresis and purified using Qiagen MinElute gel purification kit

Library strategy

ChIP-Seq

Library source

genomic

Library selection

ChIP

Instrument model

Illumina NovaSeq 6000

Data processing

Basecalls performed using RTA version 1.12.4.2
Sequenced reads passing Illumina quality filter were aligned to the Glycine max genome (Wm82.a2.v1) using Bowtie v0.12.7 with parameters -v 2 -5 0 -3 0 -m 1 --best --strata
Reads from technical replicates were combined and peaks were identified for each biological replicates using MACS3, using default parameters and a q value threshold of 0.01.
Assembly: Wm82.a2.v1
Supplementary files format and content: NarrowPeak files generated by MACS3.

Submission date

Sep 07, 2023

Last update date

Sep 16, 2023

Contact name

John Harada

E-mail(s)

[email protected]

Organization name

University of California

Department

Plant Biology