1、| | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | BRITISH STANDARD BS EN 12682:1998 The Euro
2、pean Standard EN 12682:1998 has the status of a British Standard ICS 07.080 NO COPYING WITHOUT BSI PERMISSION EXCEPT AS PERMITTED BY COPYRIGHT LAW Biotechnology Modified organisms for application in the environment Guidance for the characterization of the genetically modified organism by analysis of
3、 the functional expression of the genomic modificationBS EN 12682:1998 This British Standard, having been prepared under the direction of the Sector Committee for Materials and Chemicals, was published under the authority of the Standards Committee and comes into effect on 15 December 1998 BSI 1998
4、ISBN 0 580 30176 1 Amendments issued since publication Amd. No. Date Text affected National foreword This British Standard is the English language version of EN 12682:1998. The UK participation in its preparation was entrusted to Technical Committee CII/58, Biotechnology, which has the responsibilit
5、y to: aid enquirers to understand the text; present to the responsible European committee any enquiries on the interpretation, or proposals for change, and keep the UK interests informed; monitor related international and European developments and promulgate them in the UK. A list of organizations r
6、epresented on this committee can be obtained on request to its secretary. Cross-references The British Standards which implement international or European publications referred to in this document may be found in the BSI Standards Catalogue under the section entitled “International Standards Corresp
7、ondence Index”, or by using the “Find” facility of the BSI Standards Electronic Catalogue. A British Standard does not purport to include all the necessary provisions of a contract. Users of British Standards are responsible for their correct application. Compliance with a British Standard does not
8、of itself confer immunity from legal obligations. Summary of pages This document comprises a front cover, an inside front cover, the EN title page, pages 2 to 10, an inside back cover and a back cover.CEN European Committee for Standardization Comite Europe en de Normalisation Europa isches Komitee
9、fu r Normung Central Secretariat: rue de Stassart 36, B-1050 Brussels 1998 CEN All rights of exploitation in any form and by any means reserved worldwide for CEN national Members. Ref. No. EN 12682:1998 E EUROPEAN STANDARD EN 12682 NORME EUROPE ENNE EUROPA ISCHE NORM July 1998 ICS 07.080 Descriptors
10、: biotechnology, genetics, modified organisms, environments, environmental protection, analysis methods, bioassay, experimental design English version Biotechnology Modified organisms for application in the environment Guidance for the characterization of the genetically modified organism by analysi
11、s of the functional expression of the genomic modification Biotechnologie Organismes modifie s disse mine s dans lenvironnement Guide pour la caracte risation de lorganisme ge ne tiquement modifie par lanalyse de lexpression fonctionnelle de la modification ge nomique Biotechnik Vera nderte Organism
12、en zum Einsatz in der Umwelt Leitfaden fu r die Charakterisierung des gentechnisch vera nderten Organismus durch Untersuchung der funktionellen Auspra gung der Genomvera nderung This European Standard was approved by CEN on 1 July 1998. CEN members are bound to comply with the CEN/CENELEC Internal R
13、egulations which stipulate the conditions for giving this European Standard the status of a national standard without any alteration. Up-to-date lists and bibliographical references concerning such national standards may be obtained on application to the Central Secretariat or to any CEN member. Thi
14、s European Standard exists in three official versions (English, French, German). A version in any other language made by translation under the responsibility of a CEN member into its own language and notified to the Central Secretariat has the same status as the official versions. CEN members are th
15、e national standards bodies of Austria, Belgium, Czech Republic, Denmark, Finland, France, Germany, Greece, Iceland, Ireland, Italy, Luxembourg, Netherlands, Norway, Portugal, Spain, Sweden, Switzerland and United Kingdom.Page 2 EN 12682:1998 BSI 1998 Foreword This European Standard has been prepare
16、d by Technical Committee CEN/TC 233, Biotechnology, the Secretariat of which is held by AFNOR. This European Standard shall be given the status of a national standard, either by publication of an identical text or by endorsement, at the latest by January 1999, and conflicting national standards shal
17、l be withdrawn at the latest by January 1999. This European Standard has been prepared under a mandate given to CEN by the European Commission and the European Free Trade Association. According to the CEN/CENELEC Internal Regulations, the national standards organizations of the following countries a
18、re bound to implement this European Standard: Austria, Belgium, Czech Republic, Denmark, Finland, France, Germany, Greece, Iceland, Ireland, Italy, Luxembourg, Netherlands, Norway, Portugal, Spain, Sweden, Switzerland and the United Kingdom. Contents Page Foreword 2 Introduction 3 1 Scope 3 2 Normat
19、ive references 3 3 Definitions 3 4 Testing for functional expression 4 5 Materials 4 6 Considerations for the experimental procedures 5 7 Validity of data analysis 8 8 Documentation of results 9 Annex A (informative) Bibliography 10Page 3 EN 12682:1998 BSI 1998 Introduction This European Standard re
20、lates to the characterization of genetically modified organisms (GMOs). It is designed as a guideline for adaptation of experimental procedures to the requirements of the specific experimental design. The characterization of a GMO can include the analysis of: the genomic modification (see EN 12687);
21、 the functional expression of the genomic modification; the molecular stability of the genomic modification (see EN 12683). This European Standard deals with the analysis of the functional expression of the genomic modification of GMOs. In principle, this type of analysis can be used to correlate th
22、e functionality to the genomic modification. With respect to an intended application, it is the predicted modification of the phenotype according to the intended design which should be analysed. This European Standard deals with types of analysis performed during pre-release evaluations of GMOs. Mos
23、t types of testing performed in the laboratory are also suited for a monitoring of the GMO and its novel traits in field samples. 1 Scope This European Standard provides guidance on the design and execution of experiments for the analysis of the functional expression of the genomic modification. It
24、gives criteria for the set-up of an experimental design and the determination of the validity of its execution. The main factors influencing the specificity, reliability and limits of detection for an analysis are given for the following categories of experiments: analysis of the expression of a gen
25、omic modification within GMOs (or clones of) or any specified part of them, including its response to internal (developmental stage, growth, organ or tissue location) and external factors (temperature, humidity, pH, other); establishment of identity of a desired product resulting from the expression
26、 of a genomic modification in comparison with a reference; characterization of the novel trait(s) of a GMO. This European Standard does not cover procedures for the analysis of pleiotropic effects of a genomic modification or the act of generating the GMO (e.g. somaclonal variation) on the phenotype
27、 of modified organisms, which cannot be predicted from the knowledge of the type of the genomic modification. However, the principles stated in this European Standard are applicable for the execution of any analysis of gene functions. 2 Normative references This European Standard incorporates by dat
28、ed or undated reference, provisions from other publications. These normative references are cited at the appropriate places in the text and the publications are listed hereafter. For dated references, subsequent amendments to or revisions of any of these publications apply to this European Standard
29、only when incorporated in it by amendment or revision. For undated references, the latest edition of the publication referred to applies. EN 12687:1998, Biotechnology Modified organisms for application in the environment Guidance for the characterization of the genetically modified organism by analy
30、sis of the genomic modification. EN 12683:1998, Biotechnology Modified organisms for application in the environment Guidance for the characterization of the genetically modified organism by analysis of the molecular stability of the genomic modification. 3 Definitions For the purposes of this standa
31、rd, the following definitions apply: 3.1 analyte substance sought or determined 3.2 background noise output of a test system formed by controls lacking the data signal-giving component(s) under investigation 3.3 control preparation of known characteristics used to standardize an analysis 3.4 data si
32、gnal output of a test system NOTE Data signals can be characterized: by binary decision: presence/absence (+/2); in relative terms by ordering the data signal strength with respect to (a) defined control(s); quantitatively by giving their output strength in absolute terms; by position or movement; q
33、ualitatively by describing parameters not addressed by strength or position. 3.5 detection recognition of the presence of an organism or of a molecular structure within a sample 3.6 genetic modification of interest conceptual design for altering the genetic material within an organism NOTE 1 The gen
34、etic modification of interest can be described at different levels of molecular detail. NOTE 2 The conceptual design can include insertion, substitution or deletion of genetic material.Page 4 EN 12682:1998 BSI 1998 3.7 genetically modified organism organism in which the genetic material has been alt
35、ered in a way that does not occur naturally by mating and/or natural recombination NOTE Within the terms of this definition, genetic modification occurs at least through the use of the techniques listed in the Directive 90/220/EEC or its appropriate annexes (see annex A 2). 3.8 genomic modification
36、actual physical structure of the genetic modification of interest as it exists in the genetically modified organism 3.9 identification establishment of identity by comparison with a reference NOTE 1 The reference could be an organism, a molecular structure or the genetic modification of interest. NO
37、TE 2 The certainty of identification is affected by the types and/or number of characteristics investigated. 3.10 monitoring regular or continuous observation or collection of data with respect to an organism, process or procedure NOTE In this standard, monitoring applies to the progress of a releas
38、ed genetically modified organism. 3.11 organism biological entity capable of replication or of transferring genetic material 3.12 phenotype sum of the traits of an organism NOTE 1 The phenotype can be described with respect to one or more traits under a given set of conditions. NOTE 2 In the case of
39、 a virus, the phenotype can be described by one or more traits manifested in the infected host. 3.13 reproducibility precision under reproducibility conditions ISO 3534-1 NOTE 1 Reproducibility conditions are conditions where test results are obtained with the same method on identical test items in
40、different laboratories with different operators using different equipment. NOTE 2 Results should be expressed as reproducibility standard deviation or reproducibility coefficient of variation. 3.14 trait observable and/or measurable characteristic 4 Testing for functional expression 4.1 General cons
41、iderations The expression of traits introduced by genetic modification, over time and generation, can be important in order to ensure the performance of the GMO under field conditions and/or for biosafety reasons. The design of an analysis for functional expression should take into account the natur
42、al variations on the one hand and the experimental objective with respect to product performance and safety on the other. Data signal processing from biochemical and molecular analysis or observation of phenotypic traits is used to determine the expression caused by a genomic modification including
43、its regulation, to establish the identity of products, and to differentiate the phenotype of a GMO from that of the unmodified recipient or parent organism. 4.2 Data signal processing by methods of biochemistry, immunology and molecular biology The synthesis of gene products can be assayed by a vari
44、ety of methods (see annex A 5, 6), including: testing for sequence-dependent data signal generation of primary gene products (RNA); determination of the relative molecular mass and other physical and/or chemical characteristics of gene products; testing for the presence of specific antigenic determi
45、nants. 4.3 Phenotype testing Depending on the objective of the analysis, the characterization of a particular phenotype can efficiently substitute for a molecular analysis of the GMO such as: enzymatic activities, synthesis of metabolites; resistance or tolerance for chemicals, organisms; a new colo
46、ur or habitus phenotype; host range; behaviour. 5 Materials If nucleic acids, gene probes and labelled gene probes are used, they should be prepared in accordance with clause 5 of EN 12687:1998.Page 5 EN 12682:1998 BSI 1998 6 Considerations for the experimental procedures 6.1 General The main steps
47、of the experimental procedures for the analysis of the expression of the genomic modification are: a) precise statement of the objective for the intended analysis (see 6.2); b) experimental design according to various criteria (see 6.2); c) execution of the analysis according to the experimental pro
48、tocol (see 6.3); d) appropriate record keeping (see 6.4); e) evaluation of the validity of the results (see clause 7); f) documentation of the results (see clause 8). 6.2 Experimental design The considerations for experimental procedures to analyse the functional expression of the genetic modificati
49、on of interest should start with the definition and statement of the objective for this analysis, followed by the design of the experiment, which should be written down in a protocol, but keeping the flexibility needed to handle unexpected observations. The reason for choosing a particular method or methods should be stated in the experimental design. Gene expression is usually affected by external and internal factors influencing the organism. In some circumstances,
