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Nucleic Acids Research Advance Access originally published online on August 18, 2008
Nucleic Acids Research 2008 36(18):e118; doi:10.1093/nar/gkn524
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Nucleic Acids Research, 2008, Vol. 36, No. 18 e118
© 2008 The Author(s)
This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/2.0/uk/) which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited.

Methods Online

NAIMA: target amplification strategy allowing quantitative on-chip detection of GMOs

Dany Morisset1,*, David Dobnik1, Sandrine Hamels2, Jana Zel1 and Kristina Gruden1

1Department of Biotechnology and Systems Biology, National Institute of Biology, Vecna pot 111, Ljubljana 1000, Slovenia and 2Eppendorf Array Technologies SA, Rue du séminaire 20, B-5000 Namur, Belgium

*To whom correspondence should be addressed. Tel: +386 1 42 333 88; Fax: +386 1 25 738 47; Email: dany.morisset{at}nib.si

Received June 3, 2008. Revised July 30, 2008. Accepted July 31, 2008.

We have developed a novel multiplex quantitative DNA-based target amplification method suitable for sensitive, specific and quantitative detection on microarray. This new method named NASBA Implemented Microarray Analysis (NAIMA) was applied to GMO detection in food and feed, but its application can be extended to all fields of biology requiring simultaneous detection of low copy number DNA targets. In a first step, the use of tailed primers allows the multiplex synthesis of template DNAs in a primer extension reaction. A second step of the procedure consists of transcription-based amplification using universal primers. The cRNA product is further on directly ligated to fluorescent dyes labelled 3DNA dendrimers allowing signal amplification and hybridized without further purification on an oligonucleotide probe-based microarray for multiplex detection. Two triplex systems have been applied to test maize samples containing several transgenic lines, and NAIMA has shown to be sensitive down to two target copies and to provide quantitative data on the transgenic contents in a range of 0.1–25%. Performances of NAIMA are comparable to singleplex quantitative real-time PCR. In addition, NAIMA amplification is faster since 20 min are sufficient to achieve full amplification.


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