QRREM method for the isolation of high-quality RNA from the complex matrices of coconut.

Amjad Iqbal,Rashad Qadri,Muhammad Hamayun,Jing Li,Farooq Shah,Yaodong Yang,Anwar Hussain,Yi Wu

doi:10.1042/bsr20181163

Abstract

Complex plant tissues vary in hardness, i.e. some are succulent, while others are complex to break. Besides, plant metabolites, such as polysaccharides, proteins, polyphenols and lipids, can greatly interfere with the RNA extraction. So, in order to obtain a high-quality RNA from the complex tissues (like coconut endosperm, coconut apple and coconut leaf bud) rich in secondary metabolites, a robust method is demanded. Several methods (MRIP, CTAB and TRIZOL) have been used previously for the isolation of quality RNA from the coconut tissues, but without any success. The present study will provide with the details of a new method (Quick and Reliable RNA Extraction Method or QRREM), which have efficiently isolated the intact RNA form the complex tissues of coconut compared with CTAB, Trizol and RNA plant. The method has been validated for the isolation of high-quality intact RNA from the other available plant species (Areca/betel nut, mint and spring onion). The method has various advantages over the other methods in terms of time and cost effectiveness. Furthermore, the resulted RNA from various tissues of coconut performed well in the downstream experiments, i.e. reverse transcription and PCR for the production and amplification of cDNA.

Highlights

Many plant species contains complex tissues and higher concentrations of plant metabolites, which can interfere with the extraction of RNA
The newly developed protocol for the extraction of RNA from complex matrices, rich in carbohydrates, lipids, proteins and polyphenols provided us with a high-quality RNA in appreciable quantities when compared with the cetyltrimethylammonium bromide (CTAB), Trizol and RNAplant methods (Table 1)
The concentration of the extracted RNA from various tissues of coconut by the QRREM method was in the range from 16.2 to 63.2 μg/80 mg tissue, where for other plant species the values were between 12.7 and 20.9 (Table 2)

Summary

Introduction

Many plant species contains complex tissues and higher concentrations of plant metabolites, which can interfere with the extraction of RNA. A new method (QRREM) that gave consistent results is discussed; the method is robust, less time consuming (taking up to 1.5 h, including qualitative and quantitative analysis), cheap and reliable to isolate RNA from the very complex matrices of coconut tissues (rich in polysaccharides, lipids, proteins and polyphenols).

Results

Conclusion