Development and Validation of a Novel 29-plex STR Multiplex System for Legal and Forensic Science

Short tandem repeat (STR) analysis is the gold standard method in forensics for personal identification and paternity testing. Researchers have been working on developing novel multiplex systems containing more STRs for database construction and improvement of personal identification ability. This study's six-dye multiplex amplification system contained 29 autosomal loci, Y-indel, and Amelogenin. System optimization and performance measures were out according to the recommendations of the Scientific Working Group on DNA Analysis Methods, including PCR condition, sensitivity, mixture, inhibitor, species specificity, reproducibility, precision, stutter, concordance, and population study. The results showed that the complete profile was obtained with 125 pg of DNA input in a sensitivity study and a mixture ratio of 1:4. The full profile was observed with a hematin concentration ≤ 125 μmol/L, hemoglobin ≤ 200 μmol/L, EDTA ≤ 1.5 mmol/L, humic acid ≤ 1.5 μg/μL, indigo ≤ 12 mmol/L, and calcium ≤ 6.0 mmol/L. Meanwhile, the system also showed reasonable species specificity. Population genetic results showed the high performance of this panel with high informative and polymorphic loci, which possessed high estimates of the combined power of discrimination (1–7.16 × 10⁻³⁵) and the combined power of exclusion (1–1.98 × 10⁻¹²) in southern Han Chinese populations.