A robust plant rna isolation method suitable for affymetrix genechip analysis and quantitative real-time rt-pcr

Research output: Contribution to journalArticle

Abstract

Microarray analysis and quantitative real-time RT-PCR are the major high-throughput techniques that are used to study transcript profiles. One of the major limitations in these technologies is the isolation of large quantities of highly pure RNA from plant tissues rich in complex polysaccharides, polyphenolics and waxes. Any contamination of the isolated RNA affects the downstream applications and requires extra cleaning procedures that result in a reduced RNA yield, especially the low molecular weight molecules. The protocol presented here is suitable for isolating high yield and clean total RNA from field-grown plants. Unlike current methods, such as LiCl and TRIZOL, with this new method, the isolated RNA can be used directly for Affymetrix GeneChip labeling or real-time RT-PCR without further purification. This fast and simple protocol provides ready-to-use RNA within 4-5 h after sampling. Additionally, the protocol described here maintains the isolation of small RNA molecules, making it an ideal choice for plant RNA preparation prior to high-throughput sequencing methods to study gene expression.

Original languageEnglish (US)
Pages (from-to)333-340
Number of pages8
JournalNature Protocols
Volume4
Issue number3
DOIs
StatePublished - Mar 2009

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Plant RNA
RNA
Real-Time Polymerase Chain Reaction
Throughput
Plant Preparations
Molecules
Waxes
Microarray Analysis
Microarrays
Gene expression
Labeling
Purification
Polysaccharides
Cleaning
Contamination
Molecular Weight
Molecular weight
Tissue
Sampling
Technology

ASJC Scopus subject areas

  • Biochemistry, Genetics and Molecular Biology(all)

Cite this

A robust plant rna isolation method suitable for affymetrix genechip analysis and quantitative real-time rt-pcr. / Bilgin, Damla D.; Delucia, Evan H; Clough, Steven J.

In: Nature Protocols, Vol. 4, No. 3, 03.2009, p. 333-340.

Research output: Contribution to journalArticle

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