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Plant Science Today (PST)

Research Articles

Vol. 12 No. 3 (2025)

Optimized RNA isolation method for field-grown mature rice roots

DOI
https://doi.org/10.14719/pst.7161
Submitted
10 January 2025
Published
23-07-2025 — Updated on 31-07-2025
Versions

Abstract

High-quality RNA extraction was crucial for downstream functional genomics applications. Tissue-specific gene expression studies relied heavily on understanding molecular mechanisms at the cellular level. RNA isolation from mature field-grown rice roots posed significant challenges due to polysaccharides, secondary metabolites and other contaminants. The study contributed to development of a modified TRIzol-based RNA isolation protocol for mature field-grown rice roots. The TRIzol RNA extraction protocol was modified and compared its efficiency with four established methods: RNeasy Plant mini kit, CTAB-LiCl method, SDS method and TRIzol method. The key modifications included an increased extraction volume (1.5 mL per 100 mg tissue) and a higher ethanol concentration (80 %), along with streamlined steps to reduce processing time. The modified TRIzol method yielded an average of 537 ng/µL of high-quality RNA (A260/A280 ~ 2) from mature field-grown rice root tissues. The RNA obtained using the modified TRIzol method was validated through cDNA synthesis and amplification of the housekeeping gene actin. Syber green-based RT-qPCR revealed a single peak in the melt curve analysis, confirming the specificity of primers for cDNA binding. The optimized TRIzol protocol offered a faster, cost-effective and user-friendly approach for isolating high-quality RNA from mature field-grown rice roots compared to existing methods. Due to its high efficiency and reliability, this protocol serves as a robust and practical tool for functional genomics and molecular biology research involving challenging plant tissues.

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