The development of an 18-locus Y-STR system for forensic casework

The aim of the present work was to improve the discriminatory potential, and hence the probative value, of Y-STR-based testing by extending the set of Y chromosome STR loci available for forensic casework. In accordance with the requirements of a Y chromosome multiplex analytical system developed specifically for forensic casework use, we have sought to maximize the number of loci able to be co-amplified, ensure appropriate assay sensitivity (1–2 ng of input genomic DNA), balance inter-locus signals and minimize confounding female DNA artifacts. Two Y chromosome STR systems, multiplex I (MPI) and multiplex II (MPII), have been developed which permit the robust co-amplification of 18 Y-STRs. The loci include DYS19, DYS385(a) and (b), DYS388, DYS389I and II, DYS390, DYS391, DYS392, DYS393, DYS425, DYS434, DYS437, DYS438, DYS439, Y-GATA-C4, Y-GATA-A7.1 (DYS460) and Y-GATA-H4. The two multiplex systems are robust over a wide range of primer, magnesium, and DNA polymerase concentrations and perform well under a variety of cycling conditions. Complete male haplotypes can be obtained with as little as 100–250 pg of template DNA. Although a limited number of female DNA artifacts are observed in mixed stains in which the male DNA comprises 1/100 of the total, the male profile is easily discernible. Slightly modified versions of MPI and MPII demonstrate a significant reduction in female artifacts. Thus, it may not be necessary to employ a differential extraction strategy to obtain a male haplotype (or haplotypes in the case of multiple male donors) in cases of sexual assault. The potential utility of MPI and MPII for forensic casework is exemplified by their ability to dissect out the male haplotype in post-coital vaginal swabs and to determine the number of male donors in mixed semen stains.This study has emphasized the need for novel Y-STR multiplexes developed for forensic use to undergo a series of validation exercises that go beyond simply optimizing the PCR reaction conditions. Specifically, stringent performance checks on their efficacy need to be carried out using casework-type specimens in order to determine potential confounding effects from female DNA.     

Paternal genetic structure of Kyrgyz ethnic group in China revealed by high-resolution Y-chromosome STRs and SNPs

Kyrgyz ethnic group is one of the nomads in China, with the majority in Xinjiang and a small part of them living in Heilongjiang province. Historically, they have went through five migrations westward due to the wars. The name “Kyrgyz” means 40 tribes, originating from the primary groups of Kyrgyz. However, it is a largely understudied population, especially from the Y chromosome. In this study, we used a previously validated high-resolution Y-chromosome single nucleotide polymorphisms (Y-SNPs) and short tandem repeats (Y-STRs) system to study Kyrgyz ethnic group. A total of 314 male samples of Kyrgyz ethnic group were genotyped by 173 Y-SNPs and 27 Y-STRs. After data analysis, the results unveiled that Kyrgyz ethnic group was a population with high percentage of both haplogroup C2a1a3a1d∼-F10091 (91/134) and R1a1a1b2a2-Z2124 (109/134), which has never been reported. This implied that Kyrgyz ethnic group might have gone through bottleneck effects twice, with these two main lineages left. Mismatch analysis indicated that the biggest mismatch number in haplogroup C2a1a3a1d∼-F10091 was 10, while that of haplogroup R1a1a1b2a2-Z2124 was 20. This huge difference reflected the different substructure in two lineages, suggesting that haplogroup C2a1a3a1d∼-F10091 might have the least admixture compared to the other two lineages. After admixture modelling with other datasets, the conclusion could be drawn that Kyrgyz ethnic group had great genetic affinity with Punjabi from Lahore, Pakistan, which supported that Kyrgyz ethnic group in China was close to central Asian.     

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 1600 ng/μL humic acid). The kit exhibits high precision level with a standard deviation of allele size no more than 0.14 nt. 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.     

基于主成分分析下候选基因关联检验的数学模型

对候选基因的关联检验,都是针对性状在候选基因内使用多个SNP标记,并通过检验SNP单倍型来完成的,众所周知,多标记单倍型方法往往要比单标记方法表达出更多的信息,但是,单倍型的数量往往会随着所标记的SNP的数目增多而急剧的增加,这又会大大增加检验统计量的自由度,使用统计学中的主成分分析法来降低单倍型空间的维数,并构造关联检验来检验一个数量性状与多个单倍型的关联情况,模拟结果显示,此检验方法是较合理的.     

MPKin-YSTR: Interpretation of Y chromosome STR haplotypes for missing persons cases

Y chromosome Short Tandem Repeat (STR) haplotypes have been used in assisting forensic investigations primarily for identification and male lineage determination. The current SWGDAM interpretation guidelines for Y-STR typing provide helpful guidance on those purposes but do not address the issue of kinship analysis with Y-STR haplotypes. Because of the high mutation rate of Y-STRs, there are complex missing person cases in which inconsistent Y-STR haplotypes between true paternal lineage relatives will arise and cases with two or more male references in the same lineage and yet differ in their haplotypes. Therefore, more useful methods are needed for interpreting the Y-STR haplotype data. Computational methods and interpretation guidelines have been developed specifically addressing this issue, either using a mismatch-based counting method or a pedigree likelihood ratio method. In this study, a software program, MPKin-YSTR, was developed by implementing those more sophisticated methods. This software should be able to improve the interpretation of complex cases with Y-STR haplotype evidence. Thus, more biological evidence will be interpreted, which in turn will result in more investigation leads to help solve crimes.     

单体型推断问题与配对图

纯节俭型单体型推断(PPHI)问题是这样一类单体型推断问题给定n个基因型向量,要求寻找n对单体型,使得每一个基因型刚好由其中一对单体型组合生成,并且这2n个单体型中所含的不同单体型数目最小.u-限制单体型推断(u-PPHI)问题是一类特殊的纯节俭型单体型推断问题,要求每一个单体型至多可以用于分解u个基因型.PPHI和u-PPHI问题都是NP-困难的.文中首先介绍了配对图的概念,并通过配对图将两类问题转化为图论问题;然后分别给出了两类问题的近似算法;最后,专门讨论了当u=2时的2-PPHI问题,并在配对图上给出了相应的算法.     

基于单倍型的复杂疾病基因定位研究

本文讨论了复杂疾病基因定位研究中的单倍型关联分析方法及与之相关的单倍型推断方法,指出了目前已有方法存在的问题与面临的困难,这些困难及问题正是现代统计学研究的热点和亟待解决的问题.通过统计上的思考与理解,提出了一些有待进一步研究的问题.     

Development and validation of a new multiplex Y-STR panel designed to increase the power of discrimination

In an attempt to increase the discrimination capacity (DC) and reduce the adventitious match probability, a 6-dye multiplex Y-chromosomal short tandem repeat (Y-STR) panel named Y34plex was constructed that combined 25 Y-chromosomal markers (DYS456, DYS627, DYS390, DYS570, DYS635, DYS385a/b, DYS448, DYS437, DYS533, DYS449, DYS481, DYS392, DYS391, DYS389I, DYS460, YGATAH4, DYS438, DYS389II, DYS19, DYS458, DYF387S1a/b, DYS439, DYS393, DYS576, and DYS518) in widely used commercial kits, with nine highly polymorphic Y-STR loci (DYS557, DYS527a/b, DYS593, DYS444, DYS596, DYS643, DYS447, DYS549, and DYS645). The Y34plex is a promising type system to distinguish both unrelated and related male individuals due to the incorporation of rapidly mutated Y-STR loci. A validation study of the Y34plex was performed and followed the guidelines of the Scientific Working Group on DNA analysis methods. Results show that full Y-STR profiles were obtained from male/female DNA mixtures with 125 pg of male DNA in the presence of 50 ng of female DNA. The ability to tolerate polymerase chain reaction inhibitors commonly contained in forensic casework samples demonstrated the applicability and robustness of the Y34plex. Compared with the Yfiler Plus kit, the novel panel showed an increased power of discrimination in Chinese Wuxi Han population (n = 434). The overall haplotype diversity of the Y34plex was 0.999606, whereas DC value was 0.956221, which is suitable for use on forensic paternal investigation.     

Inference of population structure and admixture proportion from Y chromosomal data of Chinese population

In the past two decades, Y chromosome data has been generated for human population genetic studies. These Y chromosome datasets were produced with various testing methods and markers, thus difficult to combine them for a comprehensive analysis. In this study, we combine four human Y chromosomal datasets of Han, Tibetan, Hui, and Li ethnic groups. The dataset contains 27 microsatellites and 137 single nucleotide polymorphisms these populations share in common. We assembled a single dataset containing 2439 individuals from 25 nationwide populations in China. A systematic analysis of genetic distance and clustering was performed. To determine the gene flow of the studied population with worldwide populations, we modeled the ancestry informative markers. The reference panel was regarded as a mixture of South Asian (SAS), East Asian (EAS), European (EUR), African (AFR), and American (AMR) populations from 1000 Genomes data of Y chromosome using nonlinear data-fitting. We then calculated the admixture proportion of these four studied populations with 26 worldwide populations. The results showed that the Han and Hui have great genetic affinity, and Hui is the most admixed ethnic group, with 61.53% EAS, 34.65% SAS, 1.91% AFR, 1.56% AMR, and 0.04% EUR ancestry component (the AMR is highly admixed and thus should be ignored). All the other three ethnic groups contained more than 97% EAS ancestry component. The Li is the least admixed population in this study. The combined dataset in this study is the largest of this kind reported to date and proposes reference population data for use in future paternal genetic studies and forensic genealogical identification.     

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.