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.
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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
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Gehrig, H.H., Winter, K., Cushman, J. et al. An improved RNA isolation method for succulent plant species rich in polyphenols and polysaccharides. Plant Mol Biol Rep 18, 369–376 (2000). https://doi.org/10.1007/BF02825065
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DOI: https://doi.org/10.1007/BF02825065