ISO INTERNATIONAL STANDARD 21570 First edition 2005-11-01 Foodstuffs Methods of analysis for the detection of genetically modified organisms and derived products Quantitative nucleic acid based methods Produits alimentaires - Méthodes d'analyse pour la détection des organismes génétiquement modifiés et des produits derivés- Methodes quantitatives basées sur I'utilisation des acides nucleiques Reference number ISO 21570:2005(E) @ ISO 2005 Copyright International Organization for Standardization Not for Resale ISO 21570:2005(E) PDF disclaimer This PDF file may contain embedded typefaces. In accordance with Adobe's licensing policy, this file may be printed or viewed but shall not be edited unless the typefaces which are embedded are licensed to and installed on the computer performing the editing. In accepts no liability in this area. Adobe is a trademark of Adobe Systems Incorporated. Details of the software products used to create this PDF file can be found in the General Info relative to the file; the PDF-creation parameters were optimized for printing. Every care has been taken to ensure that the file is suitable for use by IsO member bodies. In ?ISO2005 All rights reserved. Unless otherwise specified, no part of this publication may be reproduced or utilized in any form or by any means, electronic or mechanical, including photocopying and microfilm, without permission in writing from either isO at the address below or IsO's member body in the country of the requester. ISO copyright office Case postale 56. CH-1211 Geneva 20 Tel. + 41 22 749 01 11 Fax + 41 22 749 09 47 E-mail [email protected] Web www.iso.org Published in Switzerland ".... CopyightInteratinal organiation for Standardization @ ISO 2005 - All rights reserved vithout license from IHS Not for Resale ISO 21570:2005(E) Contents Page Foreword. Introduction 1 Scope 2 Normative references 3 Terms and definitions. 4 Principle. 4.1 General. 4.2 Amplification, detection and confirmation of PcR products 4.3 Quantitation of PCR products .. 5 Reagents. 6 Apparatus and equipment 7 Guidelines concerning the procedure gt 7.1 General.. 7.2 Target sequence stability.. 7.3 Calibration of the analysis 3 7.4 Quantitation considerations 7.5 Quality assurance requirements 8 Interpretation. 9 Expression of results. 10 Test report.. 5 Annex A (informative) Target taxon-specific methods.. A.1 Target taxon-specific method for the absolute quantitation of the adh1 gene DNA of maize using real-time PCR... Annex B (informative) Screening methods.. .12 B.1 Screening method for the relative quantitation of the 35S-promoter DNA of soya bean line GTS 40-3-2 using real-time PCR. Annex C (informative) Construct-specific methods .. .20 C.1 Construct-specific method for the quantitation of soya bean line GTS 40-3-2 DNA using rea..time PCR (Method .... C.2 Construct-specific method for the quantitation of soya bean line GTS 40-3-2 DNA using real-time PCR (Method 2).... 27 C.3 Construct-specific method for the quantitation of Event176 maize DNA using real-time PCR...... c.4 Construct-specific method for the quantitation of soya bean line GTS 40-3-2 DNA using real-time PCR .... 41 C.5 Construct-specific method for the quantitation of maize line MON 810 DNA using real-time PCR ... .49 C.6 Construct-specific method for the quantitation of maize line Event176 DNA using real-time PCR.. .56 C.7 Construct-specific method for the quantitation of maize line Bt11 DNA using real-time PCR.... 63 ii from IHS Not for Resale ISO 21570:2005(E) C.8 Construct-specific method for the quantitation of maize line GA21 DNA using real-time PCR... C.9 Construct-specific method for the quantitation of maize line T25 DNA using real-time PCR .... 78 Annex D (informative) Event-specific methods .... D.1 Event-specific method for the absolute and relative quantitation of maize line Bt11 DNA based on real-time PcR.... .87 D.2 Event-specific method for the relative quantitation of maize line MON 810 DNA using real-time PCR... .93