Massively parallel sequencing (MPS) has rapidly become a promising method for forensic DNA typing, due to its ability to detect a large number of markers and samples simultaneously in a single reaction, and sequence information can be obtained directly. In the present study, two kinds of forensic genetic markers, short tandem repeat (STR) and identity-informative single nucleotide polymorphism (iiSNP) were analyzed simultaneously using ForenSeq DNA Signature Prep Kit, a commercially available kit on MPS platform. A total of 152 DNA markers, including 27 autosomal STR (A-STR) loci, 24 Y chromosomal STR (Y-STR) loci, 7 X chromosomal STR (X-STR) loci and 94 iiSNP loci were genotyped for 107 Tibetan individuals (53 males and 54 females). Compared with length-based STR typing methods, 112 more A-STR alleles, 41 more Y-STR alleles, and 24 more X-STR alleles were observed at 17 A-STRs, 9 Y-STRs, and 5 X-STRs using sequence-based approaches. Thirty-nine novel sequence variations were observed at 20 STR loci. When the flanking regions were also analyzed in addition to target SNPs at the 94 iiSNPs, 38 more alleles were identified. Our study provided an adequate genotype and frequencies data of the two types of genetic markers for forensic practice. Moreover, we also proved that this panel is highly polymorphic and informative in Tibetan population, and should be efficient in forensic kinship testing and personal identification cases.

Body fluids/tissue identification (BFID) is an essential procedure in forensic practice, and RNA profiling has become one of the most important methods. Small non-coding RNAs, being expressed in high copy numbers and resistant to degradation, have great potential in BFID but have not been comprehensively characterized in common forensic stains. In this study, the miRNA, piRNA, snoRNA, and snRNA were sequenced in 30 forensic relevant samples (menstrual blood, saliva, semen, skin, venous blood, and vaginal secretion) using the BGI platform. Based on small RNA profiles, relative specific markers (RSM) and absolute specific markers (ASM) were defined, which can be used to identify a specific body fluid/tissue out of two or six, respectively. A total of 5204 small RNAs were discovered including 1394 miRNAs (including 236 novel miRNA), 3157 piRNAs, 636 snoRNAs, and 17 snRNAs. RSMs for 15 pairwise body fluid/tissue groups were discovered by differential RNA analysis. In addition, 90 ASMs that were specifically expressed in a certain type of body fluid/tissue were screened, among them, snoRNAs were reported first in forensic genetics. In brief, our study deepened the understanding of small RNA profiles in forensic stains and offered potential BFID markers that can be applied in different forensic scenarios.