Yfiler Plus Research Articles

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.

A single nucleotide substitution in the flanking region of DYS533 with low peak height

We previously typed Y-STRs of 1299 Japanese samples using the Yfiler Plus kit and found three cases with a low peak of DYS533. In this study, we sequenced the flanking regions of DYS533 in these three samples and found a single nucleotide substitution at the position 111 bp upstream from the DYS533 repeat region. Because our previous study revealed that the Y-chromosomes of these three samples belong to haplogroup D-M57*, the mutation of the single nucleotide is likely to have occurred on the common ancestral Y-chromosome of these samples. The single nucleotide substitution may have caused the low peak of DYS533 in the three samples.

Developmental validation of the Yfiler Platinum PCR Amplification Kit for forensic genetic caseworks and databases.

Y chromosome kits are successfully applied in cases where human biological material exists. With the development of genotyping ability, more Y chromosomal markers are needed for finer identification of male individuals and lineages. In this study, a developmental validation of a newly emerged Y chromosome kit that combines two different kinds of markers: 38 Y-STRs and 3 Y-indels are conducted. The results show that this kit has high sensitivity when there is a small amount of DNA (125 pg), more than one male (minor:major = 1:7), or a mixture of males and females (male:female = 125pg:1875pg), inhibited substances (800 μM hematin and more than 1600ng/μL humic acid). The kit exhibits high precision level with a standard deviation of allele size no more than 0.14nt. Locus DYS481 shows the largest stutter rate, with three stutters per true allele. Population samples are well identified (MP of 0.001106), and mutations can be observed in father-son pairs (46 mutations in 70 pairs, 10 in locus DYS627). Out of all the population samples, 13.2% belong to haplogroup M117-O2a2b1a1, with their ethnic group being Han Chinese. The results show that this kit can improve the performance of identifying male individuals, obtaining more unique haplotypes (increasing from 894 to 918 of 1000 male samples) and higher discrimination capacity (increasing from 0.942 to 0.955) in this study compared to previous widely used Yfiler Plus kit. Besides, it gives information about their paternal lineages in forensic genetic casework and genealogical database construction.

Ethnic fragmentation and degree of urbanization strongly affect the discrimination power of Y-STR haplotypes in central Sahel

Y chromosome short tandem repeats (Y-STRs) are commonly used to identify male lineages for investigative and judicial purposes and could represent the only source of male-specific genetic information from unbalanced female-male mixtures. The Yfiler Plus multiplex, which includes twenty conventional and seven rapidly-mutating Y-STRs, represents the most discriminating patrilineal system commercially available to date. Over the past five years, this multiplex has been used to analyze several Eurasian populations, with a reported discrimination capacity (DC) approaching or corresponding to the highest possible value. However, despite the inclusion of rapidly mutating Y-STRs, extensive haplotype sharing was still reported for some African populations due to a number of different factors affecting the effective population size.In the present study, we analyzed 27 Y-STRs included in the Yfiler Plus multiplex and 82 Y-SNPs in central Sahel (northern Cameroon and western Chad), an African region characterized by a strong ethnic fragmentation and linguistic diversity. We evaluated the effects of population sub-structuring on genetic diversity by stratifying a sample composed of 431 males according to their ethnicity (44 different ethnic groups) and urbanization degree (four villages and four towns).Overall, we observed a low discrimination capacity (DC = 0.90), with 71 subjects (16.5 %) sharing 27 Y-STR haplotypes. Haplotype sharing was essentially limited to subjects with the same binary haplogroup, coming from the same location and belonging to the same ethnic group. Haplotype sharing was much higher in rural areas (average DC = 0.83) than urban settlements (average DC = 0.96) with a significant correlation between DC and census size (r = 0.89; p = 0.003). Notably, we found that genetic differentiation between villages from the same country (ΦST = 0.14) largely exceeded that found among countries (ΦST = 0.02). These findings have important implications for the choice of the appropriate reference population database to evaluate the statistical relevance of forensic Y-haplotype matches.

Geographical structuring and low diversity of paternal lineages in Bahrain shown by analysis of 27 Y-STRs

We have determined the distribution of Y-chromosomal haplotypes and predicted haplogroups in the ethnically diverse Kingdom of Bahrain, a small archipelago in the Arabian Gulf. Paternal population structure within Bahrain was investigated using the 27 Y-STRs (short tandem repeats) in the Yfiler Plus kit to generate haplotypes from 562 unrelated Bahraini males, sub-divided into four geographical regions—Northern, Capital, Southern and Muharraq. Yfiler Plus provided a significant improvement over the 17-locus Yfiler kit in discrimination capacity (from 77% to 87.5% overall), but discrimination capacity differed widely between regions from 98.4% in Muharraq to 75.2% in the Northern region, an unusually low value possibly resulting from recent rapid population expansion. Clusters of closely related male lineages were seen, with only 79.4% of donors displaying unique haplotypes and 59% of instances of shared haplotypes occurring within, rather than between, regions. Haplogroup prediction indicated diverse origins of the population with a predominance of haplogroups J2 and J1, both typical of the Arabian Peninsula, but also haplogroups such as B2 and E1b1a likely originating in Africa, and H, L and R2 likely indicative of migration from South Asia. Haplogroup frequencies differed significantly between regions, with J2 significantly more common in the Northern region compared with the Southern, possibly due to differential settlement by Baharna and Arabs. Our study shows that paternal lineage population structure can exist even over small geographical scales, and that highly discriminating genetic tools are required where rapid expansions have occurred within tightly bounded populations.

Haplotype analysis of 36 Y-STR loci in a Chinese Han population from Anhui Province, Eastern China.

We analyzed haplotypes for 36 Y chromosomal STRs (Y-STRs), including 27 Yfiler Plus loci and 9 additional STRs (DYS549, DYS643, DYS508, DYS447, DYS596, DYS444, DYS557, and DYS527a/b) in 2018 unrelated Chinese Han individuals from Anhui Province using DNATyperTM 36Y Kit. Phylogenetic analysis was performed to determine the genetic relationship of the Anhui Han population with other neighboring and/or linguistically close populations.

Characterizing Y-STRs in the Evaluation of Population Differentiation Using the Mean of Allele Frequency Difference between Populations.

Y-chromosomal short tandem repeats (Y-STRs) are widely used in human research for the evaluation of population substructure or population differentiation. Previous studies show that several haplotype sets can be used for the evaluation of population differentiation. However, little is known about whether each Y-STR in these sets performs well during this procedure. In this study, a total of 20,927 haplotypes of a Yfiler Plus set were collected from 41 global populations. Different configurations were observed in multidimensional scaling (MDS) plots based on pairwise genetic distances evaluated using a Yfiler set and a Yfiler Plus set, respectively. Subsequently, 23 single-copy Y-STRs were characterized in the evaluation of population differentiation using the mean of allele frequency difference (mAFD) between populations. Our results indicated that DYS392 had the largest mAFD value (0.3802) and YGATAH4 had the smallest value (0.1845). On the whole, larger pairwise genetic distances could be obtained using the set with the top fifteen markers from these 23 single-copy Y-STRs, and clear clustering or separation of populations could be observed in the MDS plot in comparison with those using the set with the minimum fifteen markers. In conclusion, the mAFD value is reliable to characterize Y-STRs for efficiency in the evaluation of population differentiation.

Y-STR chromosome structure variations: Incidence and implications

In this study 1178 Y-STR haplotypes of unrelated male donors’ samples were analyzed in order to detect Y-STR structure variations and their implications in forensic samples analysis. DNA was amplified with Yfiler or Yfiler Plus (Applied Biosystems). 19 structure variations (1.6%) were detected: 12 duplications, 5 deletions, 1 deletion/duplication and 1 complex structure variation. The markers involved were mostly DYF387S1 and DYS19. In 5 samples the structure variations included several Y-STRs. In one particular case, alleles were detected in only 7 markers of the Yfiler Plus kit. These findings are not critical for kinship studies whereas are very important in forensic caseworks interpretation.

Investigation on rapidly mutating Y-STRs multiplex in Indian population: A pilot study

Importance of rapidly mutating (RM) Y- STRs) is already established and is evident with the addition of seven RM Y-STR markers in new generation Yfiler Plus (YFP) multiplex system. Performance of the 7 RM Y-STRs included in YFP is evaluated in central Indian population. We also report here a pilot study on genetic polymorphism using 13 Rapidly Mutating Y STRs (RM Y-STR) on Indian population. Forensic parameters including allele frequencies, gene diversities, discrimination capacity and Haplotype diversity will be calculated for both the amplification assays. Forensic application and kinship analysis will be highlighted in thisstudy.

Genetic identification of Spanish civil war victims. The state of the art in Catalonia (Northeastern Spain)

At the end of 2016 and under the initiative and funding of the “Direcció General de Memòria democràtica-Departament de Justícia” (Generalitat of Catalonia), it was decided to recover and identify the remains of people disappeared in Catalonia during and after the Spanish Civil War (1936–1939).Anthropology, archaeology, history and genetics are part of the global procedure. To achieve identification by genotyping, two different genetic approaches have been carried out: 1) the directed identification through kinship analysis of alleged living relatives, when there is a previous evidence (archaeological, anthropological, historical) of a possible relationship (direct search); 2) the random crossing of a genetic profile database of victims and another database of alleged relatives in search of a possible identification (random search). The analyses of autosomal STRs, Y chromosome STRs and, in particular cases, X-InDels and mitochondrial DNA have been applied in both approaches.Here, we present the state of the process in Catalonian mass graves. Nowadays, a total of 102 post-mortem skeletal remains have been genotyped for autosomal STRs by Global Filer Express and Y-STRs from Yfiler Plus kit. Results were compared to nowadays data of living relatives in a database of 1519 genotyped for Global Filer kit, Yfiler Plus when paternally lineage was expected (248) and mitochondrial markers when maternally lineage was expected (111). Up to now, we have identified 5 victims: 4 were identified by direct strategy (including Y-STRs and mitochondrial DNA in 2 cases) and one by random search, without any previous evidence, suggesting that both strategies are useful in this context.

Single step strategy for estimating unknown STR allele frequencies in a population

Constant growth of forensic DNA databases requires new STR markers to be developed, tested and validated. Aiming to simplify the process of obtaining allele frequency distributions of new STR loci included in new megaplexes, a single step protocol was developed. This goal was achieved by pooling samples of genetically unrelated donors, previously genotyped (n = 99), containing the same amount of DNA (total DNA = 4.5 ng/ul) as measured by fluorometry. Suitability of the method was tested by amplifying commercial kits: GlobalFiler (GF), Verifiler Express (VFE) and Y-filer plus (YFP). Allele peak heights was used to estimate allele frequency and compared with the actual frequency calculated by counting method of the individual pooled samples. The multinomial goodness of fit test was performed for testing statistical differences.In case of VFE, 4.5 ng was the DNA amount that could best estimate the allele frequency distribution; 1 ng for GF and YFP. For simple, complex and compound makers, the highest allele frequencies were correctly estimated (p > 0.05) but in some cases, it was not able to detect alleles in very low frequencies, 2/198 or below, either with VFE or GF.In case of D12S391, either for VFE and GF, some microvariants with frequency of 2, 3 or 5/198 were not assigned although they were present in the profiles, probably due to POP7 resolution. In case of YFP, some alleles were not detected at DYS385.Nevertheless, with some limitations, the proposed strategy proved to be fast and efficient for major allele frequency estimation. This simple approach generates an empirical distribution that allows for the estimation of population allele frequencies in a single step assay.

LOW DISCRIMINATION POWER OF THE YFILER™ PLUS PCR AMPLIFICATION KIT IN AFRICAN POPULATIONS. DO WE NEED MORE RM Y-STRs?

Recently, thirteen rapidly mutating Y-STRs (RM Y-STRs), characterized by a mutation rate higher than 10−2/STR/generation, have been proven to be extremely useful in distinguishing among close male relatives. Six of these RM-YSTRs have been included in the Yfiler™ Plus PCR Amplification Kit, which shows the highest discriminatory power among Y-STR commercial multiplex currently available.In this study, we used Yfiler Plus to analyze 1437 males from eastern, northern and sub-Saharan Africa (462, 477 and 498 subjects, respectively). 242 out of 1437 subjects were found to share 102 Y-STR haplotypes, resulting in a low discrimination capacity (DC = 0.90) as compared to other continental regions. With few exceptions, Y-STR haplotype sharing was limited to subjects coming from the same country and ethnic group, and belonging to the same binary Y-SNP haplogroup, suggesting possible familial relationships along the male lineage.In order to evaluate the presence of hidden familial relationships, all the 242 subjects sharing a Y-STR haplotype were further genotyped for 16 autosomal STRs using the AmpFℓSTR® NGM SElect™ PCR Amplification Kit. Blind search and simulation test analyses for kinship using the Familias software revealed the presence of a low proportion of close relatives (second degree or closer) in our sample set.These findings show that close relatedness explains only a relatively small proportion of the observed Y-STR haplotype sharing, suggesting that a higher number of RM Y-STRs should be included in commercial kits to improve the discrimination power of male-specific markers in regions characterized by high levels of endogamy.

Male lineages in Brazilian populations and performance of haplogroup prediction tools

The use of Y-chromosomal genetic markers in forensic investigations demands the establishment of reliable and representative DNA databases of different reference populations. The genetic characterization of the Y chromosome variation in human populations requires the analyses of haplotype frequencies allied to haplogroup determination. The present study aimed to contribute to the Brazilian database by providing 1,382 Yfiler Plus individual profiles, from 11 Brazilian states. The Yfiler Plus markers showed high haplotype diversities in all Brazilian populations (>0.9970), allowing high intra-population discrimination in forensic investigations. Pairwise genetic distances showed a homogeneity between Brazilian populations (FST ≤ 0.0043; non-differentiation p-values ≥ 0.0212), indicating that admixed populations from Brazil can be represented in a single Yfiler Plus haplotype database, for forensic purposes. The performance of Haplogroup Predictor and NevGen software in haplogroup prediction based on Yfiler Plus and Yfiler haplotypes was evaluated in a subset of 416 Brazilian samples that were also genotyped for 51 Y-SNPs. In 25% of the samples, no classification or errors were found for at least one of the prediction tools or marker sets. NevGen presented lower error rates (5.52% and 8.65% with Yfiler Plus and Yfiler, respectively) than Haplogroup Predictor (16.11% with Yfiler Plus and 13.70% with Yfiler). In conclusion, both haplogroup prediction tools can be useful to direct the SNP typing, but present large error rates to be used in forensic analysis, especially in predicting African haplogroups in admixed South American populations.

Performance comparison of two most recent Y-STR multiplex systems for real forensic casework analysis

Detecting presence of male DNA in a mixture is still a challenge in forensic DNA typing. The study presents performance comparison of the two most recent Y-STR systems available for forensic case work, Y filer plus (YFP) and PowerPlex Y23 (PP Y23) multiplex system. Comparative study was conducted by using full and half reaction volumes of both the systems. On the basis of the work done on more than 200 case samples of sexual assault, PP Y23 multiplex system was found to be more promising than YFP. In the experiments conducted with lesser reaction volumes, it was observed that even 8 μL PCR reaction mix of PP Y23 was able to detect 16 pg of male DNA; however, with YFP it was possible with 100 pg. More balanced Y-STR profiles were observed with low male DNA with PPY23.

Y-chromosomal haplotype diversity for 27 STR loci in the Tigray population (Northern Ethiopia)

The Horn of Africa is among the main areas of origin of migrants trying to reach Europe through the so-called central Mediterranean route (from Libya to Sicily). Migration-related accidents in the Straits of Sicily are commonplace. In such circumstances, Y-STR analysis can effectively complement autosomal STR data in the identification of shipwreck victims and help reuniting families separated during the crossing.To expand currently available Y-STR haplotype reference data for Eastern Africa, the AmpFlSTR YFiler Plus PCR Amplification kit (Thermo Fisher Scientific) was used to analyze samples from 247 Ethiopian Tigray volunteer donors. The Tigray ethno-linguistic group represents over 95% of the population in the Regional state of Tigray (Northern Ethiopia), and accounts for ∼50% of the population in neighboring Eritrea.The results obtained were compared with those available for other Eastern African ethno-linguistic groups and neighbor populations from Northern Africa and the Middle East.

Is MPS always the answer? Use of two PCR-based methods for Y-chromosomal haplotyping in highly and moderately degraded bone material

Forensic and population genetics often rely on Y-chromosomal studies. Whether it is a human identification case, trace evidence examination or phylogenetic analysis, a Y-STR haplotype is an important tool in the hands of law enforcement agencies. A common obstacle in achieving satisfactory results in all of the above mentioned circumstances, is low DNA quantity and quality within samples obtained.In this study we have examined Y-STR haplotypes in 75 bone material samples, coming from different time periods. For this purpose we have chosen YFiler Plus PCR Amplification Kit (ThermoFisher Scientific) and ForenSeq Signature DNA Prep Kit (Verogen Inc.), which use two different allele calling technologies – capillary electrophoresis and Massively Parallel Sequencing respectively. Full profiles were obtained from DNA extracts with as little as 0.1896 ng (Degradation Index 1.3) (ForenSeq) and 0.0591 ng (Degradation Index 26.8) (YFiler Plus) DNA input. The results that we present in this paper show differences in amplification rates between common markers in both kits. The differences strictly reflect mean amplicon length of markers. This, however, does not seem to influence Y-haplogroup estimation results noticeably. In one sample a discordance occurred between haplotypes obtained with both methods, where a 24 allele was called in DYS390 marker by capillary electrophoresis, while for the same sample in this locus a 23 allele was shown with MPS. A reason for this is yet to be investigated. The sequence analysis revealed a significant variation between isometric alleles, especially within repetitive regions of studied Y-STR markers.

Forensic characteristics and genetic analysis of both 27 Y-STRs and 143 Y-SNPs in Eastern Han Chinese population

Y-chromosome short tandem repeat (Y-STR) and Y-chromosome single nucleotide polymorphism (Y-SNP) frequency distributions provide resources for assessment of male population stratification among world-wide populations. Currently, the Y-STR Haplotype Reference Database (YHRD) contains numerous Y-chromosome haplotype profiles from various populations and countries around the world. However, for many of the recently discovered and already phylogenetically mapped Y-SNPs, the population data are scarce. Herein, the typing of 27 Y-STRs (Yfiler Plus) and 143 Y-SNPs (self-designed Y-SNP panel) was performed on 1269 unrelated males from 11 Han Chinese populations. Haplogroup O-M175 was the most predominant haplogroup in our Han Chinese data, ranging from 67.34% (Henan Han) to 93.16% (Guangdong Han). The highest haplogroup diversity (0.967056) was observed in Heilongjiang Han, with a discrimination capacity (DC) value of 0.3723. The number of alleles at single-copy loci varied from 2 for DYS391 (Guangdong Han) to 16 for DYS518 (Henan Han). For the majority of the populations (8/11), both the haplotype diversity and DC values are 1.0000. Furthermore, genetic differentiations were observed between Northern and Southern Han Chinese. These genetic differences were mainly reflected in haplogroup distribution and frequency, and they were confirmed by statistical analysis.

Genetic structure and forensic characteristics of Tibeto-Burman-speaking \xdc-Tsang and Kham Tibetan Highlanders revealed by 27 Y-chromosomal STRs

Culturally diverse Tibetans (Ü-Tsang, Kham and Ando) harboring a unique molecular mechanism that allows them to successfully adapt to hypoxic environments in the Qinghai-Tibet Plateau have been a subject of great interest in medical genetics, linguistics, archeology and forensic science. However, forensic characteristics and genetic variations of the Y-chromosomal 27-marker haplotype included in the Yfiler Plus system in the Ü-Tsang and Kham Tibeto-Burman-speaking Tibetans remain unexplored. Thus, we genotyped 27 Y-STRs in 230 Shigatse Ü-Tsang Tibetans (SUT) and 172 Chamdo Kham Tibetans (CKT) to investigate the forensic characterization and genetic affinity of Chinese Tibetan Highlanders. The haplotype diversities were 0.999962028 in SUT and 0.999796002 in CKT. Forensic diversity measures indicated that this 27-Y-STR amplification system is appropriate for routine forensic applications, such as identifying and separating unrelated males in deficiency paternity cases, male disaster victims and missing person identification and determining male components in sexual assault cases. Moreover, the genetic relationships among 63 worldwide populations (16,282 individuals), 16 Asian populations, and 21 Chinese populations were analyzed and reconstructed using principal component analysis, multidimensional scaling plots and a phylogenetic tree. Considerable genetic differences were observed between Tibetan populations and other geographically/ethnically diverse populations (Han Chinese). Our studied SUT and CKT have a genetically closer relationship with Gansu Ando Tibetans than with other Asians. In total, our analyses indicated that subpopulation structures exist among Asian and Chinese populations, and population-specific reference databases should be established for forensic applications.

Assessment of the Yfiler® Plus PCR amplification kit for the detection of male DNA in semen-negative sexual assault cases

The ability to detect male epithelial cells deposited during digital penetration or penile penetration without ejaculation is limited by the sensitivity of the Y-STR profiling kit. In this study, the relative profiling success of the Thermofisher Yfiler® Plus kit was compared to its predecessor, AmpFlSTR Yfiler®, for 104 semen-negative sexual assault samples from casework at Forensic Science SA, Adelaide, South Australia. Yfiler Plus generated allele information in 25% more samples than Yfiler and gave a higher recovery of informative alleles in all but two samples where detectable male DNA was present. Where a profile was obtained in both kits, 92% of samples gave a higher percentage of informative loci with Yfiler Plus compared to Yfiler. Yfiler Plus also resolved DNA mixtures in 15 samples as compared to 1 sample with Yfiler. Detection of male DNA with the Quantifiler™ Trio DNA Quantification kit was shown to correlate with a successful profiling outcome with Yfiler Plus. The success of profiling with Yfiler Plus was independent of the time elapsed between the alleged offence and the sample being collected, the type of sexual penetration which occurred, and the anatomical origin of the sample.

Rapidly mutating Y-STRs in rapidly expanding populations: Discrimination power of the Yfiler Plus multiplex in northern Africa

The male-specific northern African genetic pool is characterised by a high frequency of the E-M81 haplogroup, which expanded in very recent times (2–3 kiloyears ago). As a consequence of their recent coalescence, E-M81 chromosomes often cannot be completely distinguished on the basis of their Y-STR profiles, unless rapidly-mutating Y-STRs (RM Y-STRs) are analysed. In this study, we used the Yfiler® Plus kit, which includes 7 RM Y-STRs and 20 standard Y-STR, to analyse 477 unrelated males coming from 11 northern African populations sampled from Morocco, Algeria, Libya and Egypt. The Y chromosomes were assigned to monophyletic lineages after the analysis of 72 stable biallelic polymorphisms and, as expected, we found a high proportion of E-M81 subjects (about 46%), with frequencies decreasing from west to east. We found low intra-population diversity indexes, in particular in the populations that experienced long-term isolation. The AMOVA analysis showed significant differences between the countries and between most of the 11 populations, with a rough differentiation between northwestern Africa and northeastern Africa, where the Egyptians Berbers from Siwa represented an outlier population. The comparison between the Yfiler® and the Yfiler® Plus network of the E-M81 Y chromosomes confirmed the high power of discrimination of the latter kit, thanks to higher variability of the RM Y-STRs: indeed, the number of chromosomes sharing the same haplotype was drastically reduced from 201 to 81 and limited, in the latter case, to subjects from the same population.