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Development and forensic evaluation of an 8-dye Multi-InDel multiplex panel for highly degraded samples.

Hongtao Jia1, Bowei Jiang2, Feng Wang2

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This study introduces a new 8-dye Multi-InDel panel for analyzing degraded DNA. The panel successfully genotypes samples with minimal DNA and high degradation, outperforming traditional STR kits in forensic genetics.

Keywords:
8-dye fluorescence technologyForensic geneticsHighly degraded samplesMulti-InDels

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Area of Science:

  • Forensic Genetics
  • Molecular Biology
  • Human Identification

Background:

  • Analyzing highly degraded DNA samples is a significant challenge in forensic genetics.
  • Conventional 5-dye systems have limitations in accommodating numerous loci due to short amplicon requirements and limited fluorescent channels, impacting forensic efficacy.
  • Multi-InDel markers offer advantages like high polymorphism, short amplicons, and absence of stutter, making them ideal for degraded samples.

Purpose of the Study:

  • To develop and validate an 8-dye Multi-InDel panel for enhanced forensic analysis of highly degraded DNA samples.
  • To assess the performance of the new panel in terms of DNA input requirements, thermal degradation resistance, and forensic efficiency metrics.
  • To compare the efficacy of the Multi-InDel panel against conventional Short Tandem Repeat (STR) kits for degraded sample genotyping.

Main Methods:

  • Development of an 8-dye Multi-InDel panel designed to simultaneously detect 28 autosomal Multi-InDel loci.
  • Engineering all loci to produce amplicons ≤125 bp for compatibility with degraded DNA.
  • Testing the panel's performance with low DNA input (62.5 pg) and after significant thermal degradation (100°C for 100 min).

Main Results:

  • Successful full genotyping profiles were obtained from samples with as little as 62.5 pg of DNA.
  • The 28 Multi-InDel panel maintained 100% allele detection after thermal degradation, significantly outperforming a commercial STR kit (26.2% allele detection).
  • The system demonstrated high forensic efficiency with a cumulative matching probability (CMP) of 2.3292 × 10-18 and a cumulative probability of exclusion (CPE) of 0.9999669.

Conclusions:

  • The 8-dye, 28-locus Multi-InDel panel is a robust and efficient tool for the forensic identification of highly degraded DNA samples.
  • The panel meets Scientific Working Group on DNA Analysis Methods (SWGDAM) guidelines and offers superior performance compared to conventional STR kits for challenging samples.
  • This technology provides a practical strategy for improving forensic analysis, particularly in paternity testing with degraded samples.