EN 12686-1998 en Biotechnology - Modified Organisms for Application in the Environment - Guidance for the Sampling Strategies for Deliberate Releases of Genetically Modified Micro-.pdf

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1、- STD-BSI BS EN LZbBb-ENGL 1998 Lb24bb7 0737580 Ob1 B BRITISH STANDARD Biotechnology - Modified organisms for application in the environment - Guidance for the sampling strategies for deliberate releases of genetically modified micro-organisms, including viruses The European Standard EN 126861998 ha

2、s the status of a British Standard BS EN 12686: 1998 ICs 07.080: 07.100.01 NO COPYING WITHOUT BSI PERMISSION EXCEPT AS PERMITTED BY COPYRIGHT LAW BS EN 126W.1998 direction of the Sector Committee for Materiais and Chemicals, was published under the authority of the Standards Committee and comes into

3、 effect on 16 December 1998 Amd. No. O BSi 1998 National foreword Dae Text affected This British Standard is the English language version of EN 126861998. The UK participation in its preparation was entrusted to Technical Committee CIV58, Biotechnology, which has the responsibility to: - aid enquire

4、rs to understand the text; - present to the responsible European committee any enquiries on the - monitor related international and European developments and promulgate interpretation, or proposals for change, and keep the UK interests informed, them in the UK. A iist of organizations represented on

5、 this committee can be obtained on request to ils secretary. Cross-references The Briish standards which implement international or European publications referred to in this document may be found in the BSI Standards Catalogue under the section entided “International Standards Correspondence Index“,

6、 or by using the “Find“ facility of the BSI Standards Electronic Catalogue. A British Standard does not purport to include ail the necessary provisions of a contract. Users of British Standards are responsible for their correct application. Compliance with a British Standard does not of itself confe

7、r immunity from legal obiigations. Summary of pages This document comprises a front cover, an inside front cover, the EN title page, pages 2 to 8, an inside back cover and a back cover. I I - STDDBSI BS EN 12bbb-ENGL 1998 E Zb24bb7 0739582 739 E EUROPEAN STANDARD EN 12686 NORME EUR0PEE”E EXJROP- NOR

8、M July 1998 ICs 07.080;100.01 Descripmrs: biotechnology, genetics, modified organisms, environments, environmental protection, sampling, experimental design English version Biotechnology- Mod54 organisms for application in the endment - Guidance for the samphg st;rategies for delibemte releases of g

9、enetidy modjed micrcHrganisms, includmg viruses Biotechnologie - organismes modjs cibmins dans lenvironnement - Guide des stratgies dchantillonnage pour les diss,minaons volontaires de micro-organismes gntiquement mois, y compris de Mrus Biotechnik - Vernderte Organismen zum Einsatz in der Umwelt -

10、Leitfaden fr Frobenahmestrategien bei der absichtlichen Freisetmng gentechnisch vernderter Mikroorganismen einschliefilich Viren This European Standard was approved by GEN on 1 July 1998. CEN members are bound to compiy with the CEN/CENELEC Intend Regulations which stipulate the conditions for givin

11、g this European Standard the status of a national standard without any aiteration. Uptodate lists and bibliographicd references concerning such national standards may be obtained on application to the Central Secretariat or to any CEN member. This European Standard exists in three official versions

12、(English, French, German). A version in any other language made by translation under the responsibility of a GEN member into its own language and notified to the Central Secretariat has the same status as the official versions. CEN members are the national standards bodies of Aus- Belgium, Czech Rep

13、ublic, Denmark, Finland, France, Germany, Greece, Icehd, reiand, Italy, Luxembourg, Netherlands, Norwax Portuga, Spain, Sweden, Switzerland and United Kingdom. European Committee for Standardkation Comitk Europen de Norm-on Europisches Komitee fr Normung Central Secretariat: rue de Stassart 36, B-10

14、60 Brussels O 1998 CEN All rights of exploitation in any form and by any means reserved worldwide for CEN national Members. Ref. No. EN 126861998 E Page 2 EN 12686:1998 Foreword This European Standard has been prepared by Technical Committee CENRC 233, Biotechnology, the Secretariat of which is held

15、 by AFNOR. This European Standard shall be given the status of a national standard, either by publication of an identica text or by endorsement, at the latest by January 1999, and conflicting national standards shali be withram at the latest by January 1999. This European Standard has been prepared

16、under a mandate given to CEN by the European Commission and the European Free ?kade Association. According to the CENKENELEC Internal Regulations, the national standards organizations of the following countries are bound to implement this European standard. Austria, Belgium, Czech Republic, Denmark,

17、 Fniand, France, Germany, Greece, Iceland, Ireland, I*, Luxembourg, Netheriands, Norway, Portugal, Spain, Sweden, Switzerland and the United Kingdom. Contents Foreword introduction 1 Scope 2 Normative references 3 Definitions 4 General considerations 5 samplingstrategy Annex A (informative) Relation

18、ship between a sampling and monitoring valid Annex B (informative) Bibliography pa 2 3 3 3 3 4 4 7 8 O BSI 1998 STD-BSI BS EN L2b8b-ENGL 1998 E Lb2LlbbS 073958Li 707 II Introduction When genetically modified micro-organisms including viruses (GMMs) are subject to an experimental release into the env

19、ironment, it is important to ensure the validity of sampling strategies used to monitor the release. The sampling strategy and the statistical analysis used will vary with the predicted frequency of occurrence and spatial distribution of the relevant GMMs or genes in the area studied Since there are

20、 many different techniques available for the detection and the identification of GMMS, this European Standard is formulated as a recommendation to the experimenter to design a sampling slmtegy appropriate to the purpose of the field experiment, to the micro-organisms and to the particular phenotypic

21、 and genotypic properties of the GMMs being used. This European Standard gives the experimenter a list of points that should be considered in determining the validity of a sampling slmtegy comprising valid design, review, execution and documentation of a sampling protocol. 1 Scope This European Stan

22、dard provides guidance concerning the procedures for setting up a valid sampling strategy to meet the objectives of a monitoring strategy for GMMs released into the environment. Since monitoring methods of microorganisms in environmental samples us- require pre-treatment of the samples, for example

23、the extraction and isolation of GMMs andor their nucleic acid, this is included in the scope of this European Standard. The sampling is to provide material to which subsequent analytical or biological methods for monitoring of GMMs can be applied (see EN 12685). This European Standard is intended to

24、 address sampling of micro-organisms, including vinises (and their relevant hosts), or their nucleic acid Thil, European Standard does not cover: - the sampling of vinis-like entities or similar agents; - the sampling of GMMs in food, human health and veterinary applications. NOTE Attention is drawn

25、 to national, European and international regulations, and relevant standards covering the sampling of GMMs in food, human health and veterinary applications. This European Standard can be applied to sampling of GMMs in ail habitats and micro-environments as required to meet the defined experimental

26、objectives. The mode of sampling and the nature of the samples are dependent on the particular purpose. Therefore this European Standard provides the experimenter with a list of guiding parametem that should be considered in determining the validity of the proposed strakgy for sampling. Page 3 EN 12

27、6W1998 2 Normative references This European Standard incorporates by dated 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

28、 revisions of any of these publications apply to this European Standard oniy when incorporated in it by amendment or revision. For undated references the latest edition of the publication referred to applies. EN 12685, Biotechnology - Modified organisms for applicatim in the envimment - Guidance for

29、 the mnitoriw strategies for delibemte ?deases of gmtkdy modvied micro-organimm, including wimes. 3 Denitions For the purposes of this standard, the following definitions apply: 3.1 control preparation of known characteristics used to standardize an analysis 3.2 genetically modified micro-organism m

30、icro-organism in which the genetic material has been altered 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 90/219/EEC or its appropr

31、iate annexes (see Annex B i). 3.3 genotype genetic constitution of an organism NOTE The genotype can be described with respect to particular genes. 3.4 host target species for vinis replication as defined in the experimental design 3.6 micro-environment defined location in the environment potentiall

32、y occupied by an organism NOTE This “microenvironment“ can impart a degree of confinement on the dispersal of the organism. 3.6 monitoring reguiar or continuous observation or collection of data with respect to an organism, process or procedure NOTE in this standard, monitoring applies to the progre

33、ss of a released genetically modified micro-organism. O BSI 1998 STD-BSI BS EN IZbAb-ENGL L798 Lb2qbb7 0739585 b1i3 Page 4 EN 12686:1998 3.7 monitoring strategy procedure for designjng, reviewing, executing and documenting a monitoring protocol 3.8 phenotype sum of the traits of an organism NCYE 1 T

34、he phenotype can be described with respect to one or more traits under a given set of conditions. NOTE 2 in the case of a virus, the phenotype can be descrbed by one or more traits manifested in the infected host. 3.9 release site defined area which contallis one or more experimental fields NOTE Sev

35、eral different trials can occur within a release site. 3.10 sample materials collected for analysis 3.11 sampling protocol written document describing the manner in which samples are collected, transported, stored and their pretreatment 3.12 sampling strategy procedure for designing, reviewing, exec

36、uting and documenting a sampiuig protocol 4 General considerations in order to evrlluate the validity of a proposed sampling design, the following considerations should be taken into account. The availability and use of adequate controls is essential for the validity of the results. The design of mo

37、st field triais with GMMs follows the same principles as applied to field triais with micro-organisms which are not genetically modied f3aMical analysis, required for quantitative evaluation, determines the sampling design and hence the vaiidity of the results. The purpose of the experiment should b

38、e cleariy defined in order to prepare a statistically aeqwe protocol for sampling. Specic requirements of any subsequent detection and identification method, should be considered and can influence the utility of the results for monitoring purposes (see prEN 12685). NOTE The development of a monitori

39、ng and sampling valid strategy for deliberate releases of genetically modined micro-organisms in the environment is summarized in Figure A. 1. 6 Sampling strategy 6.1 General It is nrSt necessary to determine the objectives of the sampling simtegy The main steps in the development of the sampling st

40、rategy are: a) statement of experimental objectives; b) design and review of sampling protocol; c) execution of the sampling protocol; d) appropriate record keeping. Responsibilities should be assigned for each step to a specic authority, organization or person. The sampling strategy should be revie

41、wed regulaly, for example in the light of the inspections of the release site and monitoring results, to ensure its continuing validity (see EN 12685). 6.2 Design criteria The design of the sampling protocol should meet defined objectives as determined by the experimenter. The design can vary widely

42、, given distinctive requirements for, for example, triais to study phenotypic properties of GMMs, gene transfer, or Cuspd andor persistence of GMMs in the environment. Depending on the objectives for which the sampling is carried out, the nature and composition of the samples which are collected, ca

43、n be different. Sampling can involve host materiai, soil and water, but also air (e.g. dispersai of spores), animals or plant parts (e.g. roots harbouring symbiotic micro-organisms). The protocol for sampling should be part of the preparation and design of the experiment, keeping the flexibility tha

44、t could be required by unexpected observaions, and should therefore be supported by a clear design of the experiment. Parametem such as startkg date or period, location(s) and site size, number of genotypes, are important in the development of the design. When designing the sampling protocol a very

45、important factor concerns the pretreatment of the samples which is required to allow subsequent detection and/or identification of GMMs. The choice of the pretreatment procedure is afected by the nature and composition of the samples and also by the demands of the detection andor identification tech

46、nique used for monitoring andor characterization of the GMM. Many of these techniques require extraction of micro-organisms, nucleic acid andor gene products from the sample. Examples of approaches for detection and isolation of GMMs from environmental samples are: - bioassay; - filtralion technique

47、s; - decanting and sieving; - cen-on and gradient centrifugation; - immunoiogical techniques; - mty techniques; - culturing and propagation techniques. O BSI 1998 STD-BSI BS EN 12b8b-ENGL 1778 I 1b24bb7 073758b 58T I Page 5 EN 12686:1998 For the extraction of macromolecules, such as nucleic acids an

48、d proteins, the above techniques and the following are also appropriate: - lysis by freezethawing; - extraction with organic solvent(s); - iysis using lytic enzymes; - cellular disruption techniques (e.g. disruption using -beads, disruption by sonication, disruption by a French Press); - homogenizat

49、ion. Aiternativelly, macromolecules can be extracted directly from the sample without prior isolation of a GMM. Each of the above listed ireatments generates losses of the target GMM or macromolecules. In case a combination of techniques is utilized, these losses are cumulative and can influence the results. Important points to be considered in the hitid phase in the design of the sampling protocol are. - Wd the samples be used for either qualitative or quantitative analysis? - Which statiscal method is appropriate? - Wd sampling be for