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Sample GSM5625105

Query DataSets for GSM5625105

Status

Public on Aug 30, 2023

Title

Null_Root_36DAS_LP_2

Sample type

SRA

Source name

36 days after sowing roots

Organism

Triticum aestivum

Characteristics

line: Null Segregant
age: 36 DAS
organ: Roots
treatment: Low Phosphate
cultivar: Fielder

Treatment protocol

For high phosphate (HP), the solution contained 436 µM KH2PO4, and for low phosphate (LP) this was substituted with 440 µM KCl.

Growth protocol

For evaluation of growth under low P conditions, seeds of transgenic and null segregant plants were germinated as described above, except seeds were sterilized prior to being imbibed by incubating seeds in 50% (v/v) ethanol for 15 minutes, followed by incubating in 2% (w/v) sodium perchlorate for 10 minutes, and washing 5 times with sterile ddH2O. Seeds were pre-germinated on wet filter paper on sealed petri dishes in a growth chamber at 22°C, with 16 h light (115 µE info and dark cycle for 3 days. Five to six germinated seedlings of average size and similar developmental stage were transplanted into each pot containing a mix of 70% (v/v) #20 silica sand (Gillibrand) and 30% (v/v) Green Grades Profile™. Profile™ contains 10% (v/v) aluminum oxide which acts as a solid-state buffer for phosphate77. Prior to transplanting, the growth medium of each pot was pre-washed 16 times with 0.75 L Industrial H2O, with at least 4 hours between each wash to reduce the starting phosphate content to 25 μM. For the first 14 days after sowing (DAS), plants were sub-irrigated (from pot base) with unfertilized industrial water containing negligible amounts of phosphate and nitrogen. At 14 DAS, pots were sub-irrigated with 3 L of industrial water supplemented with the following nutrients: 2.3 mM NH4NO3, 2 mM KNO3, 0.00007 mM (NH4)Mo7O24, 0.01148 mM H3BO3, 0.002 mM CuSO4, 0.0094 mM Iron-EDTA, 0.003 mM MnSO4, and 0.0035 mM ZnSO4. For high phosphate (HP), the solution contained 436 µM KH2PO4, and for low phosphate (LP) this was substituted with 440 µM KCl. The industrial water used contains 1625 µM Ca2+, 375 µM Mg2+, and 656 µM SO42-.

Extracted molecule

polyA RNA

Extraction protocol

Total RNA was extracted using TRizol reagent (Invitrogen) according to the manufacturer’s instructions. Following quantitation with a NanoDrop (Wilmington, DE), 15 µg of total-RNA was used to isolate mRNA using Dynabeads™ Oligo(dT)25 (Invitrogen) following the manufacture’s instructions.
mRNA-seq Libraries were produced using the non-strand specific Brad’s Rapid Ravi-seq procedure (Kumar et al. 2012).

Library strategy

RNA-Seq

Library source

transcriptomic

Library selection

cDNA

Instrument model

Illumina NextSeq 500

Data processing

Quality reports of raw reads were generated with the FastQC package (http://www.bioinformatics.babraham.ac.uk/projects/fastqc/).
Adapter sequences were trimmed using Cutadapt (Martin et al. 2011).
Sequences were aligned to the IWGSC v1.1 reference genome using hisat2 (Kim et al. 2015). The OsPSTOL1 K-allele CDS was added as a separate chromosome.
Read count data in terms of RPKM (reads per kilobase per million) were calculated for exons on gene ( Genomic features exonsby). For samples with both single end and paired end reads, single end and paired end reads were aligned and counted seperately as described above. The counts were then added from the two alignments
Genome_build: IWGSC v1.1
Supplementary_files_format_and_content: Normalized RPKM for each sample. P-values for comparisons are given only for genes with 5 counts per million in at least three samples.

Submission date

Oct 13, 2021

Last update date

Aug 30, 2023

Contact name

Julia Bailey-Serres

E-mail(s)

Serres@ucr.edu

Organization name

UC Riverside

Department

Department of Botany and Plant Sciences