Finally, root samples from 12 different roots per treatment group were collected for whole transcriptome analysis of wheat grown in the above-mentioned conditions. Total RNA was extracted from the collected wheat root samples using the Invitrogen Plant RNA Purification kit (Carlsbad, CA, United States). The cDNA libraries were subsequently constructed, and after quality verification, the libraries were sequenced on the Illumina HiSeq 4000 apparatus as described in our previous studies (Gong et al., 2021 (link)). The RNA-seq reads underwent filtration to eliminate any low-quality reads and adaptor sequences, with a quality score threshold of Q > 20 being applied. The wheat genome was utilized as a reference for mapping the clean reads with the assistance of the TopHat spliced read aligner (Trapnell et al., 2012 (link)). The FPKM value was utilized via Cufflinks software to assess the expression levels of mRNAs (Trapnell et al., 2010 (link)). The expression matrix of TraesTM9SF genes was then extracted from the annotated sequencing data. Next, to identify the expression profiles of TraesTM9SF genes, Short Time-series Expression Miner (STEM, http://gene.ml.cmu.edu/stem/ , (Ernst and Bar-Joseph, 2006 (link))) was run on the transcriptome data. The pheatmap package was used for visualizing the expression profile of retrieved TraesTM9SF genes.
Plant rna purification kit
Manufactured by Thermo Fisher Scientific
Sourced in United States
The Plant RNA Purification kit is a laboratory product designed to efficiently extract and purify high-quality total RNA from a variety of plant samples. The kit utilizes a reliable, column-based method to isolate RNA while removing contaminants and inhibitors.
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Lab products found in correlation
2 protocols using plant rna purification kit
1
Whole Transcriptome Analysis of Wheat Roots
2
Quantifying ABA-Responsive Gene Expression
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Example 7
To further investigate if Q27/CB, quinabactin/QB and ABA treatments induced ABA-responsive genes in vegetative tissues, qRT-PCR analysis was performed.
A plant RNA purification kit (Invitrogen) was used to isolate total RNA from plant tissue as per manufacturer's instructions. A QantiTec reverse transcription kit was used to synthesize cDNA from 1 μg of total RNA. Real-time PCR was performed using Maxima SYBR Green/Fluorescein qPCR Master Mix (Fermentas) with the iQ5 real-time PCR detection system. The relative amounts of target mRNAs were determined using the relative standard curve method and were normalized by relative amount of internal control mRNA. Biological triplicate experiments were performed.
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