Final report for CCQM-K86.b relative quantification of Bt63 in GM rice matrix sample

Abstract

A key comparison CCQM-K86.b was performed to demonstrate and document the capacity of National Metrology Institutes (NMIs) and Designated Institutes (DIs) in the determination of the relative quantity of two specific genomic DNA fragments present in a rice powder. The study provides the support for the following measurement claim: "Quantification of the ratio of the number of copies of specified intact sequence fragments of a length up to 150 nucleotides following extraction from an unprocessed, high starch ground seed matrix, with a copy number ratio from 0.005 to 1". The study was carried out under the auspices of the Nucleic Acids Working Group (NAWG) of the Consultative Committee for Amount of Substance: Metrology in Chemistry and Biology (CCQM) and was jointly coordinated by the National Institute of Metrology, China (NIMC) and Government Laboratory Hong Kong (GLHK).The following laboratories (in alphabetical order) participated in this key comparison study: CENAM (Mexico); GLHK (Hong Kong); LGC (United Kingdom); NIB (Slovenia); NIMC (P.R. of China); NIMT (Thailand); NMIA (Australia); NMIJ (Japan); TÜBITAK-UME (Turkey). Good agreement was observed between the reported results. KEY WORDS FOR SEARCH Genetically modified (GM) crops, digital PCR, copy number ratio, GM rice Main text To reach the main text of this paper, click on Final Report. Note that this text is that which appears in Appendix B of the BIPM key comparison database kcdb.bipm.org/. The final report has been peer-reviewed and approved for publication by the CCQM, according to the provisions of the CIPM Mutual Recognition Arrangement (CIPM MRA).

Consistency plots: a simple graphical tool for investigating agreement in key comparisons

“…Figure 7a) shows results and uncertainties for one of two materials examined in key comparison CCQM-K86c, a comparison on determination of a specified gene target ratio in Brassica napus (canola) [22]; for the data, see table S3 in supplementary information. The CCQM-K86 range of studies [22][23][24] supports the quantification of proportion of genetically modified (GMO) material present in otherwise wild-type materials. For this material, the nucleic acid target was a fragment introduced in modification event RT73, conferring glyphosate resistance.…”

Metrological framework to support accurate, reliable, and reproducible nucleic acid measurements

“…A series of four key comparisons enabled NMIs and DIs to demonstrate and document their capabilities in measurements of the number of copies of specified intact sequence fragments extracted from different matrices and to determine their relative quantity. In the first two studies, matrices were rich in polymeric carbohydrate (amylose and amylopectin) and poor in fat: maize (Zea maize L.) seed powder in the CCQM-K86 study [84] and rice (Oryza sativa L.) seed powder in the CCQM-K86.b study [85]. In the third study, CCQM-K86.c, high-fat/oil matrix was selected, represented by rapeseed/canola (Brassica napus L.) seed powder [86].…”