Unraveling the paternal genetic structure and forensic traits of the Hui population in Liaoning Province, China using Y-chromosome analysis.

The Hui people are the second-largest ethnic minority in China, and they are distributed throughout the country. A previous study explored the paternal genetic structure of the Hui population in nine different regions of China, but it overlooked the Liaoning province. In this study, we examined the paternal genetic makeup and forensic traits of the Hui population in Liaoning province by analyzing 157 Y-chromosome single nucleotide polymorphisms (Y-SNPs) and 26 short tandem repeats (Y-STRs). We successfully genotyped 282 unrelated male individuals from the Hui population of Liaoning province using the SNaPshot® single base extension assay and Goldeneye™ Y26 system kit (PEOPLESPOT R&D, Beijing, China). The results revealed high haplotypic diversity (0.9998) and identified 46 terminal haplogroups for the Hui population. Additional analyses, such as heat maps, principal component analysis (PCA), genetic distance (FST), Multidimensional scaling (MDS) analysis, and median-joining network (MJ) analysis, showed that the Hui population could be classified into three groups: Northwest Hui populations (NWH), including Liaoning, Xinjiang, Qinghai, Gansu, Ningxia, Shaanxi, and Henan; Hui populations from Sichuan and Shandong (SSH); and Yunnan Hui populations (YNH). Pairwise genetic distance (Rst) comparisons with other Chinese populations revealed that the Hui population displayed genetic affinity with the Han population. The comprehensive understanding of the Hui population in Liaoning province, explored by Y-SNPs and Y-STRs, can be utilized to interpret their genetic structure and enhance the accuracy of forensic databases.

Separation mixed semen of two individuals using magnetic beads coupled ABH blood group antibody.

In sexual assault cases, one of the most common samples collected is a mixed semen stain, which is often found on the vagina, female underwear, or bed sheets. However, it is usually difficult to identify the perpetrator based on this sample alone. One technique that has been developed to address this issue is magnetic bead-based separation. This method involves using modified magnetic microspheres to capture and enrich specific target cells, in this case, sperm cells. In this study, we utilized magnetic beads coupled with ABH blood group antibody to isolate sperm cells from an individual of a single ABO blood type. Subsequently, polymerase chain reaction amplification and capillary electrophoresis were employed to perform the genotyping the short tandem repeat (STR) loci. This approach allows for the identification of different individuals in a mixed seminal stain sample from two individuals, by first separating sperm cells based on ABH antigen differences and subsequently utilizing autosomal STR typing on the enriched single blood group cells.

Discrimination of monozygotic twins using mtDNA heteroplasmy through probe capture enrichment and massively parallel sequencing.

Differentiating between monozygotic (MZ) twins remains difficult because they have the same genetic makeup. Applying the traditional STR genotyping approach cannot differentiate one from the other. Heteroplasmy refers to the presence of two or more different mtDNA copies within a single cell and this phenomenon is common in humans. The levels of heteroplasmy cannot change dramatically during transmission in the female germ line but increase or decrease during germ-line transmission and in somatic tissues during life. As massively parallel sequencing (MPS) technology has advanced, it has shown the extraordinary quantity of mtDNA heteroplasmy in humans. In this study, a probe hybridization technique was used to obtain mtDNA and then MPS was performed with an average sequencing depth of above 4000. The results showed us that all ten pairs of MZ twins were clearly differentiated with the minor heteroplasmy threshold at 1.0%, 0.5%, and 0.1%, respectively. Finally, we used a probe that targeted mtDNA to boost sequencing depth without interfering with nuclear DNA and this technique can be used in forensic genetics to differentiate the MZ twins.

Large-scale pedigree analysis highlights rapidly mutating Y-chromosomal short tandem repeats for differentiating patrilineal relatives and predicting their degrees of consanguinity.

Rapidly mutating Y-chromosomal short tandem repeats (RM Y-STRs) were suggested for differentiating patrilineally related men as relevant in forensic genetics, anthropological genetics, and genetic genealogy. Empirical data are available for closely related males, while differentiation rates for more distant relatives are scarce. Available RM Y-STR mutation rate estimates are typically based on father-son pair data, while pedigree-based studies for efficient analysis requiring less samples are rare. Here, we present a large-scale pedigree analysis in 9379 pairs of men separated by 1-34 meioses on 30 Y-STRs with increased mutation rates including all known RM Y-STRs (RMplex). For comparison, part of the samples were genotyped at 25 standard Y-STRs mostly with moderate mutation rates (Yfiler Plus). For 43 of the 49 Y-STRs analyzed, pedigree-based mutation rates were similar to previous father-son based estimates, while for six markers significant differences were observed. Male relative differentiation rates from the 30 RMplex Y-STRs were 43%, 84%, 96%, 99%, and 100% for relatives separated by one, four, six, nine, and twelve meioses, respectively, which largely exceeded rates obtained by 25 standard Y-STRs. Machine learning based models for predicting the degree of patrilineal consanguinity yielded accurate and reasonably precise predictions when using RM Y-STRs. Fully matching haplotypes resulted in a 95% confidence interval of 1-6 meioses with RMplex compared to 1-25 with Yfiler Plus. Our comprehensive pedigree study demonstrates the value of RM Y-STRs for differentiating male relatives of various types, in many cases achieving individual identification, thereby overcoming the largest limitation of forensic Y-chromosome analysis.

Evaluation of facial hair-associated SNPs: a pilot study on male Pakistani Punjabi population.

Variation in facial hair is one of the most conspicuous features of facial appearance, particularly in South Asia and Middle East countries. A genome-wide association study in Latin Americans has identified multiple genetic variants at distinct loci being associated with facial hair traits including eyebrow thickness, beard thickness, and monobrow. In this pilot study, we have evaluated 16 SNPs associated with facial hair traits in 58 male individuals from the Punjabi population of Pakistan. In our sample, rs365060 in EDAR and rs12597422 in FTO showed significant association with monobrow, rs6684877 in MACF1 showed significant association with eyebrow thickness, and two SNPs in LOC105379031 (rs9654415 and rs7702331) showed significant association with beard thickness. Our results also suggest that genetic association may vary between ethnic groups and geographic regions. Although more data are needed to validate our results, our findings are of value in forensic molecular photofitting research in Pakistan.

Corrigendum: The heart of silk road "Xinjiang", its genetic portray, and forensic parameters inferred from autosomal STRs.

[This corrects the article DOI: 10.3389/fgene.2021.760760.].

Genetic diversity and forensic application of Y-filer STRs in four major ethnic groups of Pakistan.

17 Y-chromosomal STRs which are part of the Yfiler Amplification Kit were investigated in 493 unrelated Pakistani individuals belonging to the Punjabi, Sindhi, Baloch, and Pathan ethnic groups. We have assessed the forensic parameters and population genetic structure for each group. Among the 493 unrelated individuals from four ethnic groups (128 Baloch, 122 Pathan, 108 Punjabi, and 135 Sindhi), 82 haplotypes were observed with haplotype diversity (HD) of 0.9906 in Baloch, 102 haplotypes with HD value of 0.9957 in Pathans, 80 haplotypes with HD value of 0.9924 in Punjabi, and 105 haplotypes with HD value of 0.9945 in the Sindhi population. The overall gene diversity for Baloch, Pathan, Punjabi, and Sindhi populations was 0.6367, 0.6479, 0.6657, and 0.6112, respectively. The results had shown us that Pakistani populations do not have a unique set of genes but share the genetic affinity with regional (Central Asia and Northern India) populations. The observed low gene diversity (heterozygosity) values may be because of endogamy trends and this observation is equally supported by the results of forensic parameters which are mostly static across 4 combinations (minimal STRs, extended 11 Y-STRs, Powerplex 12 Y System, and Yfiler 17 Y-STRs) of STRs in these four populations.

Genetic characterization of the highlander Tibetan population from Qinghai-Tibet Plateau revealed by X chromosomal STRs.

Tibetans are considered an East Asian ethnic group and primarily live in the high Tibetan plateau, the western Sichuan and Yunnan mountains of central and southern China, and areas throughout the Himalayas and around the Tibetan plateau. These people exhibit rare molecular machinery that allows them to adapt to hypoxic environments in the Qinghai-Tibet Plateau and make them a potential candidate for providing insights related to medical genetic, molecular medicine and human population studies. In the current study, we have genotyped 549 individuals with Investigator Argus X-12 Kit. For 12 X-STRs, a total of 174 unique alleles were found, among them DXS10134 and DXS10135 were the most polymorphic loci. All of the loci were in Hardy-Weinberg Equilibrium (HWE). The numbers of observed haplotypes in Highlander Tibetans males were 161,112, 96 and 108, respectively, whereas haplotype diversities (HD) were 0.9959, 0.9880, 0.9809 and 0.9873, respectively. The combined discrimination power for males (PDm) was 0.999 999 99701 and for females (PDf) was 0.999 999 999 999 9958. This study represents an extensive report on X chromosomal STR markers variation in the Highlander Tibetans population for forensic applications and population genetic studies.

Mutation Rate Analysis of RM Y-STRs in Deep-Rooted Multi-Generational Punjabi Pedigrees from Pakistan.

Y chromosome short tandem repeat polymorphisms (Y-STRs) are important in many areas of human genetics. Y chromosomal STRs, being normally utilized in the field of forensics, exhibit low haplotype diversity in consanguineous populations and fail to discriminate among male relatives from the same pedigree. Rapidly mutating Y-STRs (RM Y-STRs) have received much attention in the past decade. These 13 RM Y-STRs have high mutation rates (>10−2) and have considerably higher haplotype diversity and discrimination capacity than conventionally used Y-STRs, showing remarkable power when it comes to differentiation in paternal lineages in endogamous populations. Previously, we analyzed two to four generations of 99 pedigrees with 1568 pairs of men covering one to six meioses from all over Pakistan and 216 male relatives from 18 deep-rooted endogamous Sindhi pedigrees covering one to seven meioses. Here, we present 861 pairs of men from 62 endogamous pedigrees covering one to six meioses from the Punjabi population of Punjab, Pakistan. Mutations were frequently observed at DYF399 and DYF403, while no mutation was observed at DYS526a/b. The rate of differentiation ranged from 29.70% (first meiosis) to 80.95% (fifth meiosis), while overall (first to sixth meiosis) differentiation was 59.46%. Combining previously published data with newly generated data, the overall differentiation rate was 38.79% based on 5176 pairs of men related by 1−20 meioses, while Yfiler differentiation was 9.24% based on 3864 pairs. Using father−son pair data from the present and previous studies, we also provide updated RM Y-STR mutation rates.

Genetic characteristics and forensic features of Xibe ethnic group revealed via extended set of Y-STRs.

BACKGROUND: Xibe is the fifth largest minority population of Liaoning province. Predominately they live in Liaoning province (69.52%), followed by Xinjiang (18.06%), Heilongjiang (3.99%), Jilin (1.63%) and Inner Mongolia provinces (1.57%). AIM: To provide an updated and precise population database on an extended set of Y STRs not available before and explore the forensic characteristics of 26 Y chromosomal STRs. SUBJECTS & METHODS: In this study, we genotyped 406 unrelated Xibe male individuals from Liaoning province using Goldeneye® 26Y System kit and calculated the forensic parameters of these 26 Y STRs loci. RESULTS: All haplotypes generated for 406 Xibe samples using Goldeneye® 26Y kit were unique with a discrimination capacity (DC) of 1. On restricting the haplotypes to the Y-filer® set of 17 Y-STRs, we observed 392 haplotypes. Among them 93.53% (380) were unique with a DC of 0.9655 and haplotype diversity (HD) of 0.9998, showing high discrimination power of the extended set of markers in this population. Allelic frequencies ranged from 0.0024 to 0.7684 across 26 Y STRs loci. DYS385 showed the highest gene diversity (0.9691) among all markers. CONCLUSION: According to pairwise RST genetic distances among Xibe populations from China, the Liaoning Xibe population showed the closest genetic distance (0.0035) followed by Xinjiang Xibe population (0.0218). Multidimensional scaling (MDS) analysis among Xibe and 29 other Chinese populations showed that local populations such as Manchu from Liaoning and Han from Beijing had a close affinity while Tibetans from Aba, China, were most distant from Xibe populations. Moreover, 12 individuals showed a null allele at DYS448 in Xibe population samples. We submitted Y-STRs data in the Y-Chromosome Haplotype Reference Database (YHRD) for future forensic and other usage.

Are Roma People Descended from the Punjab Region of Pakistan: A Y-Chromosomal Perspective.

Gypsies are a separate ethnic group living in Pakistan and some other countries as well. They are mostly known as 'Roma' and 'untouchables'. They have different types of lifestyles as compared to other common people, as they always keep migrating from one place to another. They do not have proper houses; they live in tent houses and most probably work on daily wages to earn their living. Gypsies cannot be specified according to the place of residence and can only be classified according to their migration route. Previous historical and linguistic research showed the north Indian origin of Roma people. The present study collected 285 unrelated Roma individuals living in Punjab and typed with the Goldeneye Y20 system. Allelic frequencies ranged between 0.0035 and 0.5266, with haplotype diversity (HD) of 0.9999 and discrimination capacity (DC) of 0.8790. Gene diversity (GD) ranged from 0.6489 (DYS391) to 0.9764 (DYS391) (DY385ab). A total of 223 unique alleles were observed. Interestingly, the haplogroup R accounted for 40.56% and J for 22.06%. In MDS analysis, Pakistani Roma formed a close cluster with Roma from Constanta, Romania. The migration pattern of the Roma population from Pakistan, India and Europe was inferred using coalescence theory in the Migrate-n program. Overlapping Y-STR data were used to test different migration models. These migration models showed us the dominant gene flow from Pakistan to India and Europe to Pakistan. The results of our study showed that Y STRs provided substantially stronger discriminatory power in the Pakistani Roma population.

Assessing the viability of carious lesions in human identification using STR typing.

Human teeth have become a prominent source of DNA for human forensic identification as their biological structure is highly resistant to extreme conditions. Previous forensic identification was mainly dependent on the pulp and the other hard tissues of intact teeth. However, there is high likelihood that only carious teeth can be available for forensic analysis. This study aimed to validate the use of the carious part of the teeth for forensic identification and to compare two DNA extraction methods-the operative technique with the cervical cut technique for human identification using STR typing. The reliability of STR markers in carious part of the teeth was evaluated in 120 carious teeth (60 dental pulp and 60 dentinal carious tissues, respectively) with considerable coverage of gender type and age range to avoid false exclusions. The study was performed on genuine data set where samples have been extracted by proficient dentist during the treatment operation and collected for further analysis. Complete DNA was extracted and the corresponding human identification profile was obtained using the GoldenEye™DNA ID system 20A kit. The operative technique showed a conservative approach to the sampling of carious tissues and allowed safe access to collect carious tissues, whereas the cervical cut technique permitted access to the root canals and complete sampling of the pulp tissues. The findings indicated that there was no significant association between the cervical cut and operative cut techniques (p = 0.165). In addition, there was no statistically significant association between the various teeth types and the obtained profiles observed. The operative technique, by drilling holes on the defected surface of carious human teeth and gentle hand excavation of carious tissues, was indicated to be very efficient, preserving, time-saving, and cost-effective in the recovery of human DNA from carious teeth. The result gives new insights that the carious tissues of human carious teeth might be as valid as the healthy teeth for forensic human identification.

Forensic features and genetic legacy of the Baloch population of Pakistan and the Hazara population across Durand line revealed by Y-chromosomal STRs.

The Hazara population across Durand line has experienced extensive interaction with Central Asian and East Asian populations. Hazara individuals have typical Mongolian facial appearances and they called themselves descendants of Genghis Khan's army. The people who speak the Balochi language are called Baloch. Previously, a worldwide analysis of Y-chromosomal haplotype diversity for rapidly mutating (RM) Y-STRs and with PowerPlex Y23 System (Promega Corporation Madison, USA) kit was created with collaborative efforts, but Baloch and Hazara population from Pakistan and Hazara population from Afghanistan were missing. In the current study, Yfiler Plus PCR Amplification Kit loci were examined in 260 unrelated Hazara individuals from Afghanistan, 153 Hazara individuals, and 111 Balochi individuals from Baluchistan Pakistan. For the Hazara population from Afghanistan and Pakistan overall, 380 different haplotypes were observed on these 27 Y-STR loci, gene diversities ranged from 0.51288 (DYS389I) to 0.9257 (DYF387S1), and haplotype diversity was 0.9992. For the Baloch population, every individual was unique at 27 Y-STR loci; gene diversity ranged from 0.5718 (DYS460) to 0.9371(DYF387S1). Twelve haplotypes were shared between 178 individuals, while only two haplotypes among these twelve were shared between 87 individuals in Hazara populations. Rst and Fst pairwise genetic distance analyses, multidimensional scaling plot, neighbor-joining tree, linear discriminatory analysis, and median-joining network were performed, which shed light on the history of Hazara and Baloch populations. The results of our study showed that the Yfiler Plus PCR Amplification Kit marker set provided substantially stronger discriminatory power in the Baloch population of Pakistan and the Hazara population across the Durand line.

The Heart of Silk Road "Xinjiang," Its Genetic Portray, and Forensic Parameters Inferred From Autosomal STRs.

The Xinjiang Uyghur Autonomous Region of China (XUARC) harbors almost 50 ethnic groups including the Uyghur (UGR: 45.84%), Han (HAN: 40.48%), Kazakh (KZK: 6.50%), Hui (HUI: 4.51%), Kyrgyz (KGZ: 0.86%), Mongol (MGL: 0.81%), Manchu (MCH: 0.11%), and Uzbek (UZK: 0.066%), which make it one of the most colorful regions with abundant cultural and genetic diversities. In our previous study, we established allelic frequency databases for 14 autosomal short tandem repeats (STRs) for four minority populations from XUARC (MCH, KGZ, MGL, and UZK) using the AmpFlSTR® Identifiler PCR Amplification Kit. In this study, we genotyped 2,121 samples using the GoldenEye™ 20A Kit (Beijing PeopleSpot Inc., Beijing, China) amplifying 19 autosomal STR loci for four major ethnic groups (UGR, HAN, KZK, and HUI). These groups make up 97.33% of the total XUARC population. The total number of alleles for all the 19 STRs in these populations ranged from 232 (HAN) to 224 (KZK). We did not observe any departures from the Hardy-Weinberg equilibrium (HWE) in these populations after sequential Bonferroni correction. We did find minimal departure from linkage equilibrium (LE) for a small number of pairwise combinations of loci. The match probabilities for the different populations ranged from 1 in 1.66 × 1023 (HAN) to 6.05 × 1024 (HUI), the combined power of exclusion ranged from 0.999 999 988 (HUI) to 0.999 999 993 (UGR), and the combined power of discrimination ranged from 0.999 999 999 999 999 999 999 983 (HAN) to 0.999 999 999 999 999 999 999 997 (UGR). Genetic distances, principal component analysis (PCA), STRUCTURE analysis, and the phylogenetic tree showed that genetic affinity among studied populations is consistent with linguistic, ethnic, and geographical classifications.

Tracing the Genetic Legacy of the Tibetan Empire in the Balti.

The rise and expansion of Tibetan Empire in the 7th to 9th centuries AD affected the course of history across East Eurasia, but the genetic impact of Tibetans on surrounding populations remains undefined. We sequenced 60 genomes for four populations from Pakistan and Tajikistan to explore their demographic history. We showed that the genomes of Balti people from Baltistan comprised 22.6-26% Tibetan ancestry. We inferred a single admixture event and dated it to about 39-21 generations ago, a period that postdated the conquest of Baltistan by the ancient Tibetan Empire. The analyses of mitochondrial DNA, Y, and X chromosome data indicated that both ancient Tibetan males and females were involved in the male-biased dispersal. Given the fact that the Balti people adopted Tibetan language and culture in history, our study suggested the impact of Tibetan Empire on Baltistan involved dominant cultural and minor demic diffusion.

Comprehensive biomechanical analysis of three clinically used fixation constructs for posterior malleolar fractures using cadaveric and finite element analysis.

Different fixation modalities are available for fixation of posterior malleolar fractures (PMFs), but the best method is still unclear. The purpose of this study was to carry out a comparative biomechanical analysis of three commonly used fixation constructs for PMFs using experimental and finite element analysis (FEA). 15 human cadaveric ankle specimens were randomly divided into three groups. Specimens in group-A were fixed with two anteroposterior (AP) lag screws, group-B with two posteroanterior (PA) lag screws, and for group-C, a posterior plate was used. Each model was subjected to axial load. Outcomes included loads for 0.5 mm, 1 mm, 1.5 mm, and 2 mm vertical displacements of posterior fragments were noted. 3D FE models were reconstructed from computed tomography (CT) images and subjected to vertical loads. The model's stress, fracture step-off, and resultant strains in implants were also studied in 3D FE models. Significantly higher amounts of mean compressive loads were observed to cause the same amount of vertical displacements in plate group (265 ± 60.21 N, 796 ± 57.27 N, 901.18 ± 8.88 N, 977.26 ± 13.04 N) than AP (102.7 ± 16.78 N, 169.5 ± 19.91 N, 225.32 ± 15.92 N, 269.32 ± 17.29 N) and PA (199.88 ± 31.43 N, 362.80 ± 28.46 N, 431.3 ± 28.12 N, 541.86 ± 36.05 N) lag screws respectively (P < 0.05). Simulated micro-motion analysis demonstrated that fracture step-off values in plate group (0.03 ± 0.001 mm, 0.06 ± 0.003 mm and 0.13 ± 0.010 mm) were the lowest among the three groups (P < 0.001). The cancellous bone showed the highest amount of stress in AP and PA lag groups respectively, whereas the lowest stress was noted in the plate-group. This biomechanical study concluded that posterior plating is biomechanically the most stable fixation construct for PMFs fixation. AP and PA lag screws with higher bone stress and fracture step-off values have a high tendency of bone cut-through and loss of fixation respectively.

Forensic applications and genetic characterization of Liaoning Han population revealed by extended set of autosomal STRs.

BACKGROUND: Microsatellites or short tandem repeats (STRs) are considered the gold standard for forensic investigations and autosomal STRs are used for routine forensic personal identification. AIM: To provide a precise population database on an extended set of STRs which has never been done before and explore the forensic characteristics of 20 autosomal STRs. SUBJECTS AND METHODS: In the current study, we explored the genetic characteristics of 20 STRs loci in 1138 unrelated Han individuals using Goldeneye® 20A multiplex amplification system kit in the Liaoning Han population. Additionally, phylogenetic analysis based on the Nei's standard genetic distance was performed between the Han population and other relevant populations. RESULTS: A total of 253 alleles were observed while allelic frequencies ranged from 0.00043 to 0.5369. The combined discrimination power was 99.99999999999999999999789% and the combined exclusion power was 99.999998231%. Most of the loci were in HWE while only five pairs were out of LE. Population genetic analysis showed that the Han population has similarities with other East Asian populations. CONCLUSION: GoldenEyeTM 20A kit detects high diversity in the Liaoning Han population. These STRs which are part of this kit can be used for forensic investigations. Population genetic analysis showed that the Han population is different from the minority populations of Xinjiang.