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Plant and Cell Physiology Advance Access published online on August 8, 2007
Plant and Cell Physiology, doi:10.1093/pcp/pcm103
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© The Author 2007. Published by Oxford University Press on behalf of Japanese Society of Plant Physiologists. All rights reserved. For Permissions, please e-mail: journals.permissions@oxfordjournals.org

Ultra-sensitive determination of absolute mRNA amounts at attomole levels of nearly identical plant genes with high-throughput mass spectrometry (MassARRAY)

Rust Turakulov1, Sureeporn Nontachaiyapoom2, Keith R Mitchelson1, Peter M Gresshoff2 and Artem E Men1,2

1Australian Genome Research Facility, Level 5, Gehrmann Laboratories, Research Road, University of Queensland, St. Lucia, Brisbane, 4072, Australia
2Australian Research Council Centre of Excellence for Integrative Legume Research, University of Queensland, St. Lucia, Brisbane, 4072, Australia

Corresponding author: Peter M. Gresshoff, Australian Research Council Centre of Excellence for Integrative Legume Research, University of Queensland, St. Lucia, Brisbane, 4072, Australia, Phone: 61-7-3365-3550, Fax: 61-7-3365-3550, E-mail: p.gresshoff{at}uq.edu.au


  Abstract

Detection of very small RNA amounts based on micro-dissection of plant tissue is essential for modern plant biology. Mass spectroscopy technology (MassARRAY) based on SequenomTM instrumentation was adapted to determine quickly and in a high through-put fashion (by multiplexing) the absolute amounts of mRNA of closely related soybean genes. Sensitivity of 0.1 attomoles (10-19) was achieved, representing as few as 1,000 mRNA molecules. This methodology eliminates the use of housekeeping genes as reference standards and has multiple applications for plant functional genomics, such as the monitoring of individual expression of paralogous genes at ultra-low expression levels and/or in extremely small tissue samples.

Keywords: Gene expression - multiplexing - MALDI TOF - competitive PCR - allele-specific expression


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