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Sample GSM5239821

Query DataSets for GSM5239821

Status

Public on Jun 17, 2021

Title

2mlpa-stage2-rep2

Sample type

SRA

Source name

seeds

Organism

Glycine max

Characteristics

genotype: MIPS1/mrp-l/mrp-n
tissue: seed

Treatment protocol

see growth protocol

Growth protocol

Tissue from each line was sampled at three stages of seed germination in biological triplicate with ten seeds per sample. The germination stages used were mature dry seeds (stage 1), eight hour imbibed seeds (stage 2), and germinated seeds (defined as radicle emergence; stage 3). For stage 1, seeds were ground to a fine powder using a P14 mill (Pulverisette 14, Fritsch) and stored at -80°C until use. Seeds for stages 2 and 3 were sterilized for two minutes with a 10% hypochlorite + Tris solution, washed in DI water three times for five minutes, and dried overnight. The seeds were then germinated on germination plates with filter paper and DI water in the dark at 29°C.

Extracted molecule

total RNA

Extraction protocol

Once the appropriate stage was reached, seed coats and radicles were removed, and the tissue was flash frozen with liquid nitrogen and stored at -80°C until use. The tissue from these stages was ground to powder with mortar, pestle, and liquid nitrogen.
Total RNA from all stages was extracted using the RNeasy Plant Kit with on-column DNase digestion and RLC buffer (QIAGEN, Hilden, Germany). RNA quality was determined by UV spectrophotometry (260 nm, NanoDrop1000, Thermo Fischer Scientific, Waltham, MA) and RNA integrity numbers (RIN) (BioAnalyzer, Agilent Technologies, Santa Clara, CA). Samples with a RIN value >8.0 and 260/280 ratios >2.0 were submitted to Novogene (Sacramento, CA) for mRNA sequencing. A total of 72 samples (8 lines x 3 germination stages x 3 biological replicates) were sequenced (Illumina, San Diego, CA) to acquire 30 million, 150 PE reads per sample.
paired end

Library strategy

RNA-Seq

Library source

transcriptomic

Library selection

cDNA

Instrument model

Illumina NovaSeq 6000

Description

PA_MRP_merged_samples.featurecount
D22

Data processing

Raw reads were trimmed and filtered using Skewer (version 0.2.2) to remove adapter sequences and low-quality reads and bases (<Q30)
Using STAR (version 2.5.2b), the cleaned reads were aligned to ‘Williams82,’ the well-annotated soybean reference genome (Wm82.a2.v1, downloaded from Phytozome)
Transcript abundances were calculated from the mapping results using featureCounts
These results were subsequently used for differential expression analysis with DESeq2 (version 1.22.2) in R (version 3.5.1)
Genome_build: Wm82.a2.v1, downloaded from Phytozome
Supplementary_files_format_and_content: PA_MRP_merged_samples.featurecount (represents read counts)

Submission date

Apr 13, 2021

Last update date

Jun 17, 2021

Contact name

Song Li

E-mail(s)

[email protected]

Phone

5402312756

Organization name

Virginia Tech