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Open Access Technical Note

Activated charcoal-mediated RNA extraction method for Azadirachta indica and plants highly rich in polyphenolics, polysaccharides and other complex secondary compounds

Raja Rajakani1, Lokesh Narnoliya2, Neelam Singh Sangwan2, Rajender Singh Sangwan2 and Vikrant Gupta1*

Author Affiliations

1 Biotechnology Division, CSIR-Central Institute of Medicinal and Aromatic Plants, Lucknow, 226015, India

2 Department of Metabolic and Structural Biology, CSIR-Central Institute of Medicinal and Aromatic Plants, Lucknow, 226015, India

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BMC Research Notes 2013, 6:125  doi:10.1186/1756-0500-6-125

Published: 28 March 2013

Abstract

Background

High quality RNA is a primary requisite for numerous molecular biological applications but is difficult to isolate from several plants rich in polysaccharides, polyphenolics and other secondary metabolites. These compounds either bind with nucleic acids or often co-precipitate at the final step and many times cannot be removed by conventional methods and kits. Addition of vinyl-pyrollidone polymers in extraction buffer efficiently removes polyphenolics to some extent, but, it failed in case of Azadirachta indica and several other medicinal and aromatic plants.

Findings

Here we report the use of adsorption property of activated charcoal (0.03%–0.1%) in RNA isolation procedures to remove complex secondary metabolites and polyphenolics to yield good quality RNA from Azadirachta indica. We tested and validated our modified RNA isolation method across 21 different plants including Andrographis paniculata, Aloe vera, Rosa damascena, Pelargonium graveolens, Phyllanthus amarus etc. from 13 other different families, many of which are considered as tough system for isolating RNA. The A260/280 ratio of the extracted RNA ranged between 1.8-2.0 and distinct 28S and 18S ribosomal RNA bands were observed in denaturing agarose gel electrophoresis. Analysis using Agilent 2100 Bioanalyzer revealed intact total RNA yield with very good RNA Integrity Number.

Conclusions

The RNA isolated by our modified method was found to be of high quality and amenable for sensitive downstream molecular applications like subtractive library construction and RT-PCR. This modified RNA isolation procedure would aid and accelerate the biotechnological studies in complex medicinal and aromatic plants which are extremely rich in secondary metabolic compounds.

Keywords:
Activated charcoal; CTAB; Medicinal plants; Polysaccharides; RNA isolation; Secondary metabolite