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Extraction and electrophoretic analysis of large dsRNAs from desiccated plant tissues infected with plant viruses and biotrophic fungi

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Abstract

Extraction and electrophoretic analysis of large dsRNAs from plant and fungal tissues has been used successfully to detect RNA viruses infecting plants, fungi, and oomycetes. We modified a previously reported dsRNA extraction protocol and used it to detect a wide variety of plant and putative fungal viruses from infected plant tissues. The modified protocol was used successfully to extract large dsRNAs from 50 to 70 mg of desiccated plant tissues infected with acute and persistent RNA viruses and from plant tissues infected with biotrophic fungi causing rusts and powdery mildew diseases. The protocol proved to be efficient, fast, economic and versatile and requires relatively small amounts of desiccated tissue.

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Abbreviations

BaEV :

Basella alba endornavirus

BPEV :

Bell pepper endornavirus

BMV :

Brome mosaic virus

BYDV-PAV :

Barley yellow dwarf virus-PAV

CTV :

Citrus tristeza virus

DsRNA :

Double-stranded ribonucleic acid

ELISA :

Enzyme-linked immunosorbent assay

EtOH :

Ethanol

JHFMoV :

Japanese holly fern mottle virus

PCV1 :

Pepper cryptic virus 1

PCV2 :

Pepper cryptic virus 2

PcV :

Penicillium chrysogenum virus

PMV :

Panicum mosaic virus

PMMoV :

Pepper mild mottle virus

PvEV1 :

Phaseolus vulgaris endornavirus 1

RT-PCR :

Reverse transcription polymerase chain reaction

SMV :

Soybean mosaic virus

NaAc :

Sodium acetate

STV :

Southern tomato virus

STE :

Sodium chloride tris EDTA

TMV :

Tomato mosaic virus

TMGMV :

Tomato mild green mosaic virus

TAE :

Tris-acetate EDTA

TNV :

Tobacco necrosis virus

TRSV :

Tobacco ringspot virus

UmV-H1 :

Ustilago maydis virus H1

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Acknowledgments

The authors wish to thank Andrea Hebert, Middleton Library, Louisiana State University for providing valuable suggestions to the manuscript. This research was partially supported by funds from the National Institute of Food and Agriculture, the Louisiana Soybean and Grain Research and Promotion Board, and a grant to E. Rodrigues de Souto from Coordenacăo de Aperfeicoamento de Pessoal de Nível Superior (CAPES), Brasil.

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Authors and Affiliations

  1. Department of Plant Pathology and Crop Physiology, Louisiana State University Agricultural Center, Baton Rouge, LA, 70803, USA

    S. Khankhum, C. Escalante & R. A. Valverde

  2. Departamento de Agronomia, Universidade Estadual de Maringa, Parana, Maringa, 87020-900, Brazil

    E. Rodrigues de Souto and

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  1. S. Khankhum
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  2. C. Escalante
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Correspondence to R. A. Valverde.

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Khankhum, S., Escalante, C., de Souto and, E.R. et al. Extraction and electrophoretic analysis of large dsRNAs from desiccated plant tissues infected with plant viruses and biotrophic fungi. Eur J Plant Pathol 147, 431–441 (2017). https://doi.org/10.1007/s10658-016-1014-7

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