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Development of a multiplex methylation-sensitive restriction enzyme-based SNP typing system for deconvolution of semen-containing mixtures

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Abstract

The identification of mixed stains has always been a difficult problem in personal identification in the forensic field. In recent years, tissue-specific methylation sites have proven to be very stable biomarkers for distinguishing tissue origin. However, it is still challenging to perform tissue source identification and individual identification simultaneously. In this study, we developed a method that uses tissue-specific methylation markers combined with single-nucleotide polymorphism (SNP) markers to detect semen from mixed biofluids and to identify individuals simultaneously. Semen-specific CpG markers were chosen from the literature and further validated utilizing methylation-sensitive restriction endonuclease (MSRE) combined with PCR technology. The neighboring SNP markers were searched in the flanking sequence of the target CpG within 400 bp, and SNP typing was then carried out through a single-base extension reaction followed by capillary electrophoresis. Eventually, a method of MSRE combined with SNaPshot that could detect 12 compound CpG-SNP markers was developed. Using this system, 10 ng of total DNA and DNA mixture with semen content up to 25% could be typed successfully. Moreover, the cumulative discrimination power of the system in the northern Chinese Han population is 0.9998. This study provides a valuable strategy for forensic practice to perform tissue origin and individual identification from mixed stains simultaneously.

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All data generated or analyzed during this study are included in this published article and its supplementary materials.

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Funding

This work was supported by the National Natural Science Foundation of China [No.81701868 and No.82030058], the Startup Foundation for Doctors of Shanxi Medical University [BS03201621], and the Key Research and Development (R&D) Projects of Shanxi Province [No.201803D31069].

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Author notes

  1. Zeqin Li and Jintao Li have contributed equally for this work.

Authors and Affiliations

  1. School of Forensic Medicine, Shanxi Medical University, Jinzhong, Shanxi, 030619, People’s Republic of China

    Zeqin Li, Jintao Li, Yidan Li, Na Liu, Feng Liu, Jianbo Ren, Keming Yun, Jiangwei Yan & Gengqian Zhang

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  1. Zeqin Li
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  2. Jintao Li
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  3. Yidan Li
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  4. Na Liu
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  6. Jianbo Ren
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  7. Keming Yun
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  8. Jiangwei Yan
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Contributions

The authors Gengqian Zhang and Zeqin Li contributed to the study conception and design. Material preparation, data collection, and analysis were performed by Zeqin Li, Jintao Li, Yidan Li, Na Liu, Feng Liu, and Jianbo Ren. The first draft of the manuscript was written by Zeqin Li and Jintao Li, and all authors commented on previous versions of the manuscript. The authors Jiangwei Yan, Gengqian Zhang, and Keming Yun revised it critically for important intellectual content. All authors read and approved the final manuscript.

Corresponding authors

Correspondence to Keming Yun, Jiangwei Yan or Gengqian Zhang.

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Ethics approval

Approval was obtained from the ethics committee of Shanxi Medical University (No. 2019sll014). The procedures used in this study adhere to the tenets of the Declaration of Helsinki.

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Informed consent was obtained from all individual participants included in the study.

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The authors affirm that human research participants provided informed consent for publication of the data in Supplementary Table 6.

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Li, Z., Li, J., Li, Y. et al. Development of a multiplex methylation-sensitive restriction enzyme-based SNP typing system for deconvolution of semen-containing mixtures. Int J Legal Med 135, 1281–1294 (2021). https://doi.org/10.1007/s00414-021-02552-9

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  • DOI: https://doi.org/10.1007/s00414-021-02552-9

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