BS ISO 22753:2021 pdf download – Molecular biomarker analysis — Method for the statistical evaluation of analytical results obtained in testing sub-sampled groups of genetically modified seeds and grains — General requirements
The process of forming seed/grain groups from a series of sampling steps starting with the bulk seed/grain lot is shown in Figure 1, (1). Although the procedures for obtaining a laboratory sample from a seed/grain lot is not the subject of this document, a laboratory sample (2) from a seed/grain lot shall be obtained appropriately.
The procedures can be designed according to the References , , , , , ,  and . The laboratory sample shall be thoroughly mixed and divided/reduced to create the test sample (3). Likewise, the test sample shall be thoroughly mixed (i.e. homogeneous) and divided into seed/grain groups (each group represented as an array in Figure 1, (4)) following simple random sampling principles. The seed/grain groups can vary in size from one single seed/grain up to the complete test sample (i.e. a single bulk).
In most cases, multiple seed/grain groups are created from the test sample.
A determined number of seeds/grains can either be obtained by weighing or a volumetric measurement, where an approximation of number is made based on a determined conversion factor (e.g. thousand seeds/grains weight). For the case that weight is used to obtain the seed/grain groups, the operator shall have an estimate of the variability introduced by using weight rather than seed/grain count. The group testing procedure described in Clause 7 is carried out on the collective qualitative (positive or negative) results for each seed/grain group.
4.3 Detection methods for the qualitative analysis of GM seed/grain in seed/grain groups In general, GMO detection methods are categorized into two classes [ 21 ] .
The first class of assays targets a nucleic acid sequence for detecting GMO presence. The second class includes methods for detecting a specified protein that confers a specific transgenic trait. Detection methods from either or both classes should be selected considering fitness-for-purpose. Guidance on the selection of qualitative methods is provided in Clause 8. Further details can be found in ISO 21569 [ 4 ] and ISO 21572.
4.4 Statistical evaluation
Sampling and measurement uncertainty shall be considered. Sampling uncertainty can be adequately considered using the binomial distribution [ 18 ][ 2 ] .
The FPR and the FNR of the qualitative assay should be considered [ 2 ] . The LOD of the applied detection method should be considered. The group testing described here can be used to set reject/accept criteria based on a given threshold by GMO content, as well as to estimate the GMO content and associated upper and lower confidence limits.
All reagents used in the analysis should be those specified in the method. Otherwise, all reagents should be of molecular biology grade. These reagents shall be stored and used as recommended by the supplier or according to the laboratory quality assurance specifications. It can also be appropriate to aliquot the reaction solutions required for the analytical method in order to avoid subjecting them to repeated freeze–thaw cycles, or to reduce the chances of cross contamination or both. Further details shall refer to ISO 24276 and ISO 21572.
6 Apparatus and equipment
The laboratory should use properly maintained equipment suitable for the methods employed.
Further details shall refer to ISO 24276 and ISO 21572.
7 Design of testing plan
The number of seeds/grains tested, the reject/accept criteria, the sample preparation steps and the method used for testing shall be determined depending on the analytical purpose.
In seed/grain sample classification, it can be determined whether the number of deviant seeds/grains or seed/grain groups is above a given reject/accept criterion or not. Then, it can be decided to reject or accept the seed/grain lot based on the test results.
A basic testing plan for group testing consists of three fundamental parameters:
a) the number of seed/grain groups;
b) the size of the seed/grain groups;BS ISO 22753 pdf download.