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Multicolor Real-Time PCR Genotyping of ABO System Using Displacing Probes

Authors:	Li Ruan PhD ; Huian Zhao BS ; and Qingge Li PhD

Institution:	Molecular Diagnostics Laboratory, The Key Laboratory of The Ministry of Education for Cell Biology and Tumor Cell Engineering, Department of Biomedicine, School of Life Sciences, Xiamen University, Xiamen, Fujian 361005, China.; Department of Public Security of Jiangxi Province, Institute of Forensic Sciences, Nanchang, Jiangxi 330006, China.; Center of Translational Medicine, Institute for Biomedical Research, Xiamen University, Xiamen, Fujian 361005, China.

Abstract:	Abstract: Rapid and informative ABO genotyping has become increasingly popular in forensic use. We developed a multiplex real-time polymerase chain reaction (PCR) approach to genotype ABO major groups and subgroups. Seven differently fluorophor-labeled displacing probes for O¹(261delG), A(261G), A(796C/803C), B(796A/803C), O² (802G>A), A² (1059delC), and A² (1009A>G) were combined in one or two PCRs to determine either ABO major groups or subgroups. The method correctly detected 13 reference DNA samples. A blind test of 237 samples resulted in complete agreement with their phenotypes, and 110 of these 237 samples as well as with PCR-SSP method. The whole analysis could be finished in less than 100 min at substantially low material cost and the template DNA ranging from 0.16 to 500 ng per reaction could be quantitatively detected. Despite the limited informativeness of ABO genotyping, the developed methods could find application in rapid and inexpensive screening of forensic settings.

Keywords:	forensic science genotyping ABO blood group real-time polymerase chain reaction displacing probes single nucleotide polymorphisms

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