Optimized SDS-Based Protocol for High-Quality RNA Extraction from Musa spp

Area of Science:

• Plant Molecular Biology
• Biotechnology
• Biochemistry

Background:

• Banana (Musa spp.) tissues contain high levels of polyphenols and polysaccharides, hindering nucleic acid extraction.
• Previous RNA extraction methods for Musa spp. often result in low yield and poor quality RNA.

Purpose of the Study:

• To develop and validate a modified SDS-based RNA extraction method for Musa spp.
• To improve RNA yield and quality from various banana tissues, including drought-stressed samples.
• To ensure the extracted RNA is suitable for downstream molecular applications.

Main Methods:

• Modification of the RNA extraction buffer composition, including SDS concentration and the addition of LiCl.
• Inclusion of a heat incubation step during the extraction process.
• Assessment of RNA quality and yield using gel electrophoresis, spectrophotometry (A260/280 and A260/230 ratios), and RNA Integrity Number (RNA IQ).

Main Results:

• The modified method significantly increased RNA yield and quality across different Musa varieties and tissue types.
• High-quality RNA was obtained, evidenced by clear ribosomal RNA bands, absence of genomic DNA contamination, and optimal A260/280 (1.83–2.25) and A260/230 ratios.
• RNA extracted from drought-stressed plants showed high integrity (RNA IQ 7.8–9.9) and yield (2.92–6.30 µg/100 mg fresh weight).
• Successfully applied in Polymerase Chain Reaction (PCR) and quantitative real-time PCR (qRT-PCR) for gene expression analysis.

Conclusions:

• The modified SDS-based method is a reliable and efficient technique for extracting high-quality RNA from Musa spp.
• This method surpasses the limitations of traditional RNA extraction procedures like Trizol, NucleoSpin, RNeasy, and CTAB for banana tissues.
• The improved RNA extraction protocol facilitates accurate gene expression studies in banana.