Exploitation of a novel slowly mutating Y-STRs set and evaluation of slowly mutating Y-STRs plus Y-SNPs typing strategy in forensic genetics and evolutionary research.

The Y-chromosome short tandem repeats (Y-STRs) loci with different mutation rates existing in the Y chromosome non-recombination region (NRY) allow to be applied in human forensics, genealogical researches, historical investigations and evolutionary studies. Currently, there is a high demand for pedigree search to narrow the scope of crime investigations. However, the commonly used Y-STRs kits generally contain Y-STRs with high mutation rates that could cause individuals from the same pedigree to display different haplotypes. Herein, we put forward a new strategy of Slowly Mutating (SM) Y-STRs plus Y-SNPs typing, which could not only improve the resolution and accuracy of pedigree search, but also be applicable to evolutionary research. First, we developed a nine SM Y-STRs assay by evaluating their mutation rates in 210 pedigrees. Then the gene diversity and efficiency of the SM Y-STRs and 172 Y-SNPs sets were investigated by 2304 unrelated males from 24 populations. Furthermore, network and time estimation analyses were performed to evaluate the new strategy's capability to reconstruct phylogenetic tree and reliability to infer the time to the most recent common ancestor (TMRCA). The nine SM Y-STRs assay even had a higher resolution and a comparable capacity of revealing population genetic differentiation compared to 172 Y-SNPs system. This new strategy could optimize the phylogenetic tree generated by commonly used Y-STR panels and obtain a quite consistent time estimations with the published dating.

Expression profile analysis and stability evaluation of 18 small RNAs in the Chinese Han population.

In recent years, differentially expressed small RNAs have been widely used to identify the compositions of forensically relevant biological samples, and a vast number of such RNA candidates have been proposed. Nevertheless, when assessing the expression levels of target small RNAs using relative quantitative analysis methods, credible internal controls are usually required for reliable data normalization. Therefore, the identification of optimal reference genes is an important task. In this study, the expression profile of 18 small RNA reference genes was characterized in the Chinese Han population using TaqMan real-time quantitative PCR. Systematic evaluations of these candidate genes were performed based on their expression levels and stability in several common types of body fluids (i.e., venous blood, menstrual blood, saliva, semen, and vaginal secretions). Analysis results from the ΔCq method, BestKeeper, NormFinder, and geNorm were integrated by RefFinder for ranking and comparing the candidates in each type of body fluid. Among all the candidates, miR-191 was identified as the most suitable reference gene because it had a favorable ranking value in all tested samples. In addition, miR-423, miR-93, miR-484, and let-7i were also shown to be applicable reference genes. Overall, this study provides detailed assessment results of these candidate genes in different body fluids; thus, it could be used as a guide for the selection of reference genes according to their performance in the sample of choice.

Genetic insights into the paternal admixture history of Chinese Mongolians via high-resolution customized Y-SNP SNaPshot panels.

The Mongolian people, one of the Mongolic-speaking populations, are native to the Mongolian Plateau in North China and southern Siberia. Many ancient DNA studies recently reported extensive population transformations during the Paleolithic to historic periods in this region, while little is known about the paternal genetic legacy of modern geographically different Mongolians. Here, we genotyped 215 Y-chromosomal single nucleotide polymorphisms (Y-SNPs) and 37 Y-chromosomal short tandem repeats (Y-STRs) among 679 Mongolian individuals from Hohhot, Hulunbuir, and Ordos in North China using the AGCU Y37 kit and our developed eight Y-SNP SNaPshot panels (including two panels first reported herein). The C-M130 Y-SNP SNaPshot panel defines 28 subhaplogroups, and the N/O/Q complementary Y-SNP SNaPshot panel defines 30 subhaplogroups of N1b-F2930, N1a1a1a1a3-B197, Q-M242, and O2a2b1a1a1a4a-CTS4658, which improved the resolution our developed Y-SNP SNaPshot panel set and could be applied for dissecting the finer-scale paternal lineages of Mongolic speakers. We found a strong association between Mongolian-prevailing haplogroups and some observed microvariants among the newly generated Y-STR haplotype data, suggesting the possibility of haplogroup prediction based on the distribution of Y-STR haplotypes. We identified three main ancestral sources of the observed Mongolian-dominant haplogroups, including the local lineage of C2*-M217 and incoming lineages from other regions of southern East Asia (O2*-M122, O1b*-P31, and N1*-CTS3750) and western Eurasia (R1*-M173). We also observed DE-M145, D1*-M174, C1*-F3393, G*-M201, I-M170, J*-M304, L-M20, O1a*-M119, and Q*-M242 at relatively low frequencies (< 5.00%), suggesting a complex admixture history between Mongolians and other incoming Eurasians from surrounding regions. Genetic clustering analyses indicated that the studied Mongolians showed close genetic affinities with other Altaic-speaking populations and Sinitic-speaking Hui people. The Y-SNP haplotype/haplogroup-based genetic legacy not only revealed that the stratification among geographically/linguistically/ethnically different Chinese populations was highly consistent with the geographical division and language classification, but also demonstrated that patrilineal genetic materials could provide fine-scale genetic structures among geographically different Mongolian people, suggesting that our developed high-resolution Y-SNP SNaPshot panels have the potential for forensic pedigree searches and biogeographical ancestry inference.

Massively parallel sequencing of 165 ancestry-informative SNPs and forensic biogeographical ancestry inference in three southern Chinese Sinitic/Tai-Kadai populations.

Ancestry informative markers (AIMs), which are distributed throughout the human genome, harbor significant allele frequency differences among diverse ethnic groups. The use of sets of AIMs to reconstruct population history and genetic relationships is attracting interest in the forensic community, because biogeographic ancestry information for a casework sample can potentially be predicted and used to guide the investigative process. However, subpopulation ancestry inference within East Asia remains in its infancy due to a lack of population reference data collection and incomplete validation work on newly developed or commercial AIM sets. In the present study, 316 Chinese persons, including 85 Sinitic-speaking Haikou Han, 120 Qiongzhong Hlai and 111 Daozhen Gelao individuals belonging to Tai-Kadai-speaking populations, were analyzed using the Precision ID Ancestry Panel (165 AISNPs). Combined with our previous 165-AISNP data (375 individuals from 6 populations), the 1000 Genomes Project and forensic literature, comprehensive population genetic comparisons and ancestry inference were further performed via ADMIXTURE, TreeMix, PCA, f-statistics and N-J tree. Although several nonpolymorphic loci were identified in the three southern Chinese populations, the forensic parameters of this ancestry inference panel were better than those for the 23 STR-based Huaxia Platinum System, which is suitable for use as a robust tool in forensic individual identification and parentage testing. The results based on the ancestry assignment and admixture proportion evaluation revealed that this panel could be used successfully to assign individuals at a continental scale but also possessed obvious limitations in discriminatory power in intercontinental individuals, especially for European-Asian admixed Uyghurs or in populations lacking reference databases. Population genetic analyses further revealed five continental population clusters and three East Asian-focused population subgroups, which is consistent with linguistic affiliations. Ancestry composition and multiple phylogenetic analysis further demonstrated that the geographically isolated Qiongzhong Hlai harbored a close phylogenetic relationship with Austronesian speakers and possessed a homogenous Tai-Kadai-dominant ancestry, which could be used as the ancestral source proxy in population history reconstruction of Tai-Kadai-speaking populations and as one of the representatives for forensic database establishment. In summary, more population-specific AIM sets focused on East Asian subpopulations, comprehensive algorithms and high-coverage population reference data should be developed and validated in the next step.