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Nucleic acid extraction and virus detection in grapevine.

M A Rezaian¹, L R Krake

¹CSIRO, Division of Horticultural Research, Adelaide, S.A., Australia.

Journal of Virological Methods

|September 1, 1987

Summary

Grapevine nucleic acid extraction is challenging due to polyphenols. A rapid sodium perchlorate method efficiently isolates high and low molecular weight RNA, enabling sensitive viral RNA detection.

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Area of Science:

Plant molecular biology
Biochemistry
Virology

Background:

Grapevine tissues are rich in polyphenols and other compounds that inhibit nucleic acid extraction.
Conventional methods using phenol or salt/detergent-based media yield poor results, with low RNA yield or degradation.

Purpose of the Study:

To compare different extraction media for isolating nucleic acids from grapevines.
To identify an efficient method for obtaining high-quality RNA from polyphenol-rich grapevine tissues.
To enable the detection of viral RNA in grapevines.

Main Methods:

Comparison of three nucleic acid extraction media: phenol-containing, conventional salt/detergent-based, and sodium perchlorate.
Assessment of RNA yield and integrity (high and low molecular weight RNA).
Dot-blot hybridization for viral RNA detection.

Main Results:

Phenol-containing media yielded small amounts of soluble RNA and no high molecular weight ribosomal RNA (rRNA).
Salt/detergent media extracted rRNA, but significant RNA breakdown occurred.
Sodium perchlorate rapidly and efficiently extracted both high and low molecular weight RNA.
Viral RNA, undetectable with phenol extraction, was detected at picogram levels using the sodium perchlorate method.

Conclusions:

Sodium perchlorate is a superior chaotropic salt for rapid and efficient nucleic acid extraction from grapevines.
This method overcomes the inhibitory effects of polyphenols and allows for sensitive detection of viral RNA.
The developed procedure is crucial for grapevine research and diagnostics, particularly for viral disease identification.

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