Interlaboratory validation data on real-time polymerase chain reaction detection for unauthorized genetically modified papaya line PRSV-YK

This article is referred to research article entitled “Whole genome sequence analysis of unidentified genetically modified papaya for development of a specific detection method” (Nakamura et al., 2016) [1]. Real-time polymerase chain reaction (PCR) detection method for unauthorized genetically modified (GM) papaya (Carica papaya L.) line PRSV-YK (PRSV-YK detection method) was developed using whole genome sequence data (DDBJ Sequenced Read Archive under accession No. PRJDB3976). Interlaboratory validation datasets for PRSV-YK detection method were provided. Data indicating homogeneity of samples prepared for interlaboratory validation were included. Specificity and sensitivity test data for PRSV-YK detection method were also provided.


Value of the data
The data support development of real-time PCR detection method for GM papaya using whole genome sequence data.
The data provide information on reliability of developed real-time PCR method to detect GM papaya line PRSV-YK.
The data support developed real-time PCR method use in monitoring foods for GM papaya line PRSV-YK contamination.

Data
Datasets provided in this article represent reliability of unauthorized genetically modified (GM) papaya (Carica papaya L.) line PRSV-YK detection method (PRSV-YK detection method), including papaya endogenous gene, Chymopapain (Chy), detection method, using real-time polymerase chain reaction (PCR). Table 1 presents specificity of PRSV-YK and Chy detection methods. Fig. 1 shows that Chy detection method amplified papaya DNA, but both PRSV-YK and Chy detection methods did not amplify rice, soybean, maize, potato, rapeseed, pineapple, peach or passion fruit DNA. Fig. 2 presents sensitivity of PRSV-YK detection method. Cycle threshold (Ct) values obtained from real-time PCR amplification plot were quantitative (R 2 ¼0.99) in the range of 0.01-100% line PRSV-YK DNA concentrations. Table 2 presents results of homogeneity test on prepared samples. Table 3 presents statistical data obtained from homogeneity test on prepared samples. Table 4 summarizes interlaboratory validation data. Data were statistically analyzed to determine mean, relative standard deviation (RSD), repeatability RSD (RSD r ) and reproducibility RSD (RSD R ) from Ct values obtained [1].

Preparation of samples for interlaboratory validation
DNA purified from fresh papaya fruit was used as sample. DNA purified from GM papaya was mixed with DNA from non-GM papaya to prepare a dilution series of GM papaya DNA. Samples were prepared at three different levels of GM papaya DNA concentrations (0%, 0.05% and 0.10% [w/w]). Aliquots of the diluted DNA were placed in individual tubes. Each tube was then labeled with a randomized number. Six randomly selected tubes of each analyte concentration were tested as blind a Ct values (threshold value at 0.2) were recorded from duplicate test using DNA purified from each sample. b À , scored negative.
samples at each participating laboratory owning an ABI PRISM 7900HT Sequence Detection System (Thermo Fisher Scientific Inc.). Samples and real-time PCR primer and probe solutions were stored frozen at À 20°C until use.

Confirmation of homogeneity of samples
According to a procedure described by Thompson et al. [2], homogeneity of samples was verified before dispatching them to participating laboratories. Ten test samples of each GM papaya DNA   concentration (0%, 0.05% and 0.10% [w/w]) were labeled with a randomized number, and randomly selected samples were used. Each blind sample was tested to determine Ct values at threshold value 0.2 from exponential amplification plots obtained using developed real-time PCR method [1]. Data were analyzed by Cochran's test and one-way analysis of variance.

Interlaboratory validation
Method for interlaboratory validation was followed as described previously [2,3]. Twelve laboratory participants were organized to evaluate repeatability and reproducibility of developed real-time PCR method. Reagents and accessories necessary for real-time PCR and experimental protocol were provided to each participating laboratory. ABI PRISM 7900HT Sequence Detection System, owned by each lab, was used for analyses. Real-time PCR was conducted within three months. All data were collected from 12 laboratories. Presence of line PRSV-YK in samples was judged by Ct values at threshold value 0.2 (present, Ct o48.00; absent, Ct Z48.00) obtained using PRSV-YK and Chy detection methods. To statistically analyze interlaboratory validation data, Ct values from all laboratories were used after eliminating outliers by a 1-tailed Cochran's test at a probability value of 2.5%.