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Sample GSM5795756

Query DataSets for GSM5795756

Status

Public on Jan 13, 2022

Title

sparse panicle1 mutant inflorescence_12DAS_rep2

Sample type

SRA

Source name

early_inflorescence

Organism

Setaria viridis

Characteristics

genotype/variation: sparse panicle1 mutant
developmental stage: 12DAS

Growth protocol

Mutants in the sparse panicle1 locus (spp1) were used in this study. Seeds were planted in MetroMix 360 soil (Sun Gro Horticulture company) and grown in a controlled growth chamber with the following conditions: temperature of 31°C/23°C (day/night), light intensity of 200 µmol/sq.meter/sec for 12 hours (6am-6pm) and 50% relative humidity, at the Donald Danforth Plant Science Center’s Integrated Plant Growth Facility. Plants were fertilized with Jack’s 15-16-17 (Hummert International) twice a week.

Extracted molecule

total RNA

Extraction protocol

Inflorescences were hand-dissected into fresh 100% acetone on dry ice from wildtype (A10.1) and mutant (sparse panicle1) S. viridis seedlings at 10, 12, and 14DAS. Depending on the representative size at a given stage, 10-30 individual inflorescence primordia were pooled per biological replicate; three to four biological replicates were collected for each stage. All sampling was performed within a two-hour window in the morning to control for circadian effects. Acetone was removed and samples were flash-frozen in liquid nitrogen and ground into a fine powder using 3mm tungsten-carbide beads (Qiagen) in a Tissue Lyser-II. Total RNA was isolated using the PicoPure RNA Isolation Kit (Thermo Fisher Scientific) with in-column DNase I treatment following manufacturer's protocols.
RNA-seq libraries were generated from 1µg total RNA using the NEBNext Ultra Directional RNA Library Prep Kit for Illumina (New England BioLabs Inc.) and size-selected for 200 bp inserts. Libraries were quantified on a bioanalyzer (Agilent 2100) using a DNA 1000 chip to confirm the insert size and quantified again on a Qubit 3.0 Fluorometer (Life Technologies) to ensure different libraries were equally loaded
Libraries were sequenced using standard Illumina protocols (Illumina, Inc.) for either paired-end (PE) sequencing at 100bp on the Illumina HiSeq 2500 platform at the University of Illinois at Urbana-Champaign W.M. Keck Center.

Library strategy

RNA-Seq

Library source

transcriptomic

Library selection

cDNA

Instrument model

Illumina HiSeq 2500

Description

spp1_12DAS_R2

Data processing

Adaptors and low-quality reads were trimmed using Trimmomatic (Bolger et al., 2014) and reads were quality-checked using fastqc after trimming.
The S. viridis reference genome (v1) was indexed using bowtie 2 (Langmead and Salzberg, 2012) from Sviridis_311_v1.0.fa.gz file at PhytozomeV11 (phytozome.jgi.doe.gov). Reads were mapped to the reference genome using tophat2 and differentially expressed genes were identified using cuffdiff (Trapnell et al., 2012).
Expression levels quantified in Fragments Per Kilobase of transcript per Million (FPKM) were extracted for 35,214 S. viridis primary transcripts (Tables S5, S6). Gene annotation and grass homolog identification followed Zhu et al. ((2018)).
Genome_build: Genome_build: Sviridis_311_v1
Supplementary_files_format_and_content: Processed data files are MS Excel spreadsheets. Table S5. RNA-seq library sequencing and mapping statistics. Table S6. Expression of all S. viridis genes from each replicate (R1-R4) of the developmental stages (10, 12, 14 DAS) in A10.1 and spp1. Table S7. Expression of differentially expressed genes between A10.1 and spp1 at each developmental stage (10, 12, 14 DAS).

Submission date

Jan 10, 2022

Last update date

Jan 13, 2022

Contact name

Elizabeth A. Kellogg

E-mail(s)

ekellogg@danforthcenter.org

Phone

3145871490

Organization name

Donald Danforth Plant Science Center