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Updated: Jun 11, 2026
Single Droplet Digital Polymerase Chain Reaction for Comprehensive and Simultaneous Detection of Mutations in Hotspot Regions
Published on: September 25, 2018
Simultaneous detection of trisomies 13, 18, and 21 with multiplex ligation-dependent probe amplification-based
Qiwei Guo1, Yulin Zhou, Xiaobo Wang
1Engineering Research Centre of Molecular Diagnostics, Ministry of Education, Department of Biomedical Sciences and the Key Laboratory of the Ministry of Education for Cell Biology and Tumor Cell Engineering, School of Life Sciences, Xiamen University, Xiamen, China.
Background:
Trisomies 13, 18, and 21 account for the majority of chromosomal aneuploidies detected in prenatal diagnosis. Diagnosis of these trisomies relies mainly on karyotype analysis. Several molecular methods have been developed for trisomy detection, but performance or throughput limitations of these methods currently constrain their use in routine testing.
Methods:
We developed multiplex ligation-dependent probe amplification-based real-time PCR (MLPA/rtPCR) to simultaneously detect these 3 trisomy conditions with a single reaction. We applied the method to DNA isolated from 144 blinded clinical samples that included 32 cases of trisomy 21, 11 cases of trisomy 18, 1 case of trisomy 13, and 100 unaffected control samples; results were compared with karyotype analysis.
Results:
As judged by the results of the karyotype analysis, MLPA/rtPCR correctly detected all 44 cases of trisomy in the analysis of the blinded clinical samples. The method was able to detect a change in chromosome dosage as low as 1.2-fold.
Conclusions:
This novel PCR-based technology simultaneously identified 3 types of trisomy in a single reaction and accurately detected trisomy with mosaicism, while reducing assay times and costs compared with conventional methods. The MLPA/rtPCR approach may have applicability in noninvasive prenatal diagnosis with maternal blood samples.
