Plant Molecular Biology Reporter

, Volume 18, Issue 4, pp 369–376

An improved RNA isolation method for succulent plant species rich in polyphenols and polysaccharides

  • H. H. Gehrig
  • K. Winter
  • J. Cushman
  • A. Borland
  • T. Taybi
Protocols

DOI: 10.1007/BF02825065

Cite this article as:
Gehrig, H.H., Winter, K., Cushman, J. et al. Plant Mol Biol Rep (2000) 18: 369. doi:10.1007/BF02825065

Abstract

Isolation of high quality, high molecular weight RNA from plant tissues is complicated by high levels of polyphenols and polysaccharides which bind to and/or co-precipitate with RNA. Using high molecular weight polyethylene glycol (HMW-PEG), RNA was successfully isolated from plant species in which other RNA extraction methods and commercially available kits failed to deliver suitable results. We tested various buffer systems and isolation conditions with and without PEG or PVP (polyvinylpyrrolidone) using tissue from species ofAloe, Ananas, Clusia, Euphorbia, Kalanchoe, Opuntia, andPyrrosia, all of which contain high amounts of phenolic compounds and/or polysaccharides. HMW-PEG was found to be more effective than PVP in removing these compounds. RNA extraction using HMW-PEG resulted in RNA of high quality from all species investigated, as indicated by UV light absorption profiles, and also yielded PCR amplification products after reverse transcription.

Key words

AloeAnanasClusiacrassulacean acid metabolismEuphorbiaKalanchoeOpuntiaphenolic compoundsPyrrosiaRNA extraction

Abbreviations

CAM

crassulacean acid metabolism

DEPC

diethylpyrocarbonate

EtBr

ethidium bromide

FW

fresh weight

GHCL

guanidinium hydrochloride

GITC

guanidinium isothiocyanate

HMW-PEG

high-molecular-weight polyethylene glycol

NaAc

sodium-acetate

PCI

phenol/chloroform/isoamyl-alcohol-mix

PVP

polyvinylpyrrolidone

RT

room temperature

RT-PCR

reverse-transcription-polymerase chain reaction

TRIR

Trizol-reagent

UV

ultraviolet

Copyright information

© Springer 2000

Authors and Affiliations

  • H. H. Gehrig
    • 1
  • K. Winter
    • 1
  • J. Cushman
    • 2
  • A. Borland
    • 3
  • T. Taybi
    • 3
  1. 1.Smithsonian Tropical Research InstituteBalboaRepublic of Panama
  2. 2.University of Nevada, MS200RenoUSA
  3. 3.Dept. of Agricultural and Environmental SciencesUniversity of NewcastleNewcastle upon TyneU.K.