Limits...
A highly sensitive and specific method for the screening detection of genetically modified organisms based on digital PCR without pretreatment.

Fu W, Zhu P, Wang C, Huang K, Du Z, Tian W, Wang Q, Wang H, Xu W, Zhu S - Sci Rep (2015)

Bottom Line: However, to use this improved method, the samples must be pretreated, which could introduce inaccuracy into the results.The results showed that the limit of detection of our method was 0.1%, which was lower than the labeling threshold level of the EU.In conclusion, the method in our study would allow more sensitive, specific and stable screening detection of the GMO content of international trading products.

View Article: PubMed Central - PubMed

Affiliation: The Institute of Plant Quarantine, Chinese Academy of Inspection and Quarantine, Beijing, 100029, China.

ABSTRACT
Digital PCR has developed rapidly since it was first reported in the 1990 s. It was recently reported that an improved method facilitated the detection of genetically modified organisms (GMOs). However, to use this improved method, the samples must be pretreated, which could introduce inaccuracy into the results. In our study, we explored a pretreatment-free digital PCR detection method for the screening for GMOs. We chose the CaMV35s promoter and the NOS terminator as the templates in our assay. To determine the specificity of our method, 9 events of GMOs were collected, including MON810, MON863, TC1507, MIR604, MIR162, GA21, T25, NK603 and Bt176. Moreover, the sensitivity, intra-laboratory and inter-laboratory reproducibility of our detection method were assessed. The results showed that the limit of detection of our method was 0.1%, which was lower than the labeling threshold level of the EU. The specificity and stability among the 9 events were consistent, respectively. The intra-laboratory and inter-laboratory reproducibility were both good. Finally, the perfect fitness for the detection of eight double-blind samples indicated the good practicability of our method. In conclusion, the method in our study would allow more sensitive, specific and stable screening detection of the GMO content of international trading products.

No MeSH data available.


The linear curve between the theoretical copy number of transgenes and copy number of the screening elements.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC4530665&req=5

f3: The linear curve between the theoretical copy number of transgenes and copy number of the screening elements.

Mentions: The commercial platforms for conducting dPCR mainly involve two systems: the chamber-based digital PCR (cdPCR) and the droplet-based digital PCR (ddPCR) system. The cdPCR system is based on microfluidic chambers into which the reaction volume is partitioned, whereas the ddPCR system is based on droplets of water-in-oil emulsion. The different principles at play in these two systems may result in different detection results. We designed experiments to ascertain that our detection method was suited to these other dPCR platforms. The verification results are shown in Fig. 3. The results obtained using the ddPCR system indicated that the detection method we developed in this study had good inter-platform repeatability.


A highly sensitive and specific method for the screening detection of genetically modified organisms based on digital PCR without pretreatment.

Fu W, Zhu P, Wang C, Huang K, Du Z, Tian W, Wang Q, Wang H, Xu W, Zhu S - Sci Rep (2015)

The linear curve between the theoretical copy number of transgenes and copy number of the screening elements.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4530665&req=5

f3: The linear curve between the theoretical copy number of transgenes and copy number of the screening elements.
Mentions: The commercial platforms for conducting dPCR mainly involve two systems: the chamber-based digital PCR (cdPCR) and the droplet-based digital PCR (ddPCR) system. The cdPCR system is based on microfluidic chambers into which the reaction volume is partitioned, whereas the ddPCR system is based on droplets of water-in-oil emulsion. The different principles at play in these two systems may result in different detection results. We designed experiments to ascertain that our detection method was suited to these other dPCR platforms. The verification results are shown in Fig. 3. The results obtained using the ddPCR system indicated that the detection method we developed in this study had good inter-platform repeatability.

Bottom Line: However, to use this improved method, the samples must be pretreated, which could introduce inaccuracy into the results.The results showed that the limit of detection of our method was 0.1%, which was lower than the labeling threshold level of the EU.In conclusion, the method in our study would allow more sensitive, specific and stable screening detection of the GMO content of international trading products.

View Article: PubMed Central - PubMed

Affiliation: The Institute of Plant Quarantine, Chinese Academy of Inspection and Quarantine, Beijing, 100029, China.

ABSTRACT
Digital PCR has developed rapidly since it was first reported in the 1990 s. It was recently reported that an improved method facilitated the detection of genetically modified organisms (GMOs). However, to use this improved method, the samples must be pretreated, which could introduce inaccuracy into the results. In our study, we explored a pretreatment-free digital PCR detection method for the screening for GMOs. We chose the CaMV35s promoter and the NOS terminator as the templates in our assay. To determine the specificity of our method, 9 events of GMOs were collected, including MON810, MON863, TC1507, MIR604, MIR162, GA21, T25, NK603 and Bt176. Moreover, the sensitivity, intra-laboratory and inter-laboratory reproducibility of our detection method were assessed. The results showed that the limit of detection of our method was 0.1%, which was lower than the labeling threshold level of the EU. The specificity and stability among the 9 events were consistent, respectively. The intra-laboratory and inter-laboratory reproducibility were both good. Finally, the perfect fitness for the detection of eight double-blind samples indicated the good practicability of our method. In conclusion, the method in our study would allow more sensitive, specific and stable screening detection of the GMO content of international trading products.

No MeSH data available.