Population Genetics Research Research Articles

A Reevaluation of the Status of <i>Lilium nepalense</i> Based on Complete Chloroplast Genome Sequences

Lilium nepalense D. Don is an ornamental plant and exhibits high morphological variation across its distribution range in China. There is incongruence between different data sets for the status of it in Lilium. In this study, we reported a complete chloroplast genome of L. nepalense sampled in Hengduan Mountains, China. The whole chloroplast genome possessed a total length of 152,206 bp with typical circular structure, and contained a large single copy (LSC) of 81,854 bp and a small single copy (SSC) of 17,563 bp, which were separated by a pair of inverted repeats (IRa/IRb) of 26,399 bp. The average GC content among the whole chloroplast genome sequence was 37%, and the GC content in LSC, SSC, IRs regions were 34.8%, 30.6%, and 42.5%, respectively. There were 135 genes detected from the whole chloroplast genome sequence, including 89 protein-coding genes, 38 tRNAs, and 8 rRNAs. Phylogenetic results using maximum likelihood and Bayesian methods showed congruent results that L. nepalense together with the other two L. nepalense samples collected from different habitats formed a single branch, indicating a close relationship of L. nepalense with L. taliense belonging to the section Sinomartagon. This newly characterized chloroplast genome will provide essential data for the further population genetics research of L. nepalense.

Overcoming constraints on the detection of recessive selection in human genes from population frequency data

The identification of genes that evolve under recessive natural selection is a long-standing goal of population genetics research that has important applications to the discovery of genes associated with disease. We found that commonly used methods to evaluate selective constraint at the gene level are highly sensitive to genes under heterozygous selection but ubiquitously fail to detect recessively evolving genes. Additionally, more sophisticated likelihood-based methods designed to detect recessivity similarly lack power for a human gene of realistic length from current population sample sizes. However, extensive simulations suggested that recessive genes may be detectable in aggregate. Here, we offer a method informed by population genetics simulations designed to detect recessive purifying selection in gene sets. Applying this to empirical gene sets produced significant enrichments for strong recessive selection in genes previously inferred to be under recessive selection in a consanguineous cohort and in genes involved in autosomal recessive monogenic disorders.

Waste not, want not: Microsatellites remain an economical and informative technology for conservation genetics.

Comparisons of microsatellites and single‐nucleotide polymorphisms (SNPs) have found that SNPs outperform microsatellites in population genetic analyses, questioning the continued utility of microsatellites in population and landscape genetics. Yet, highly polymorphic markers may be of value in species that have reduced genetic variation. This study repeated previous analyses that used microsatellites with SNPs developed from ddRAD sequencing in the black‐capped vireo source‐sink system. SNPs provided greater resolution of genetic diversity, population differentiation, and migrant detection but could not reconstruct parentage relationships due to insufficient heterozygosities. The biological inferences made by both sets of markers were similar: asymmetrical gene flow from source sites to the remaining sink sites. With the landscape genetic analyses, we found different results between the two molecular markers, but associations of the top environmental features (riparian, open habitat, agriculture, and human development) with dispersal estimates were shared between marker types. Despite the higher precision of SNPs, we find that microsatellites effectively uncover population processes and patterns and are superior for parentage analyses in this species with reduced genetic diversity. This study illustrates the continued applicability and relevance of microsatellites in population genetic research.

The complete mitochondrial genome of Artemia salina Leach, 1819 (Crustacea: Anostraca)

In the study, the complete mitochondrial genome of Artemia salina was reported for the first time. The mitochondrial genome of A. salina is 15,762 bp in length, with the typical structure of 13 protein-coding genes (PCGs), 22 transfer RNA genes (tRNAs), and two ribosomal RNA genes, and a major non-coding region (CR). Phylogenetic analysis showed that A. salina has a much closer relationship with A. persimilis compared to other Artemia species. The complete cp genome sequence of A. salina reported here provided an essential resource for further population genetics research and germplasm conservation on Artemia.

Genotypes of Rhizophora Propagules From a Non-mangrove Beach Provide Evidence of Recent Long-Distance Dispersal

Dispersal plays a crucial role in the connectivity of established mangrove populations and in species range dynamics. As species ranges shift in response to climate change, range expansions can occur from incremental short-distance dispersal events and from stochastic long-distance dispersal events. Most population genetic research dealt with historically accumulated events though evidence of actual propagule dispersal allows to estimate genotypic features and origin of founders. In this study, we aim to disentangle a contemporary dispersal event. Using microsatellite markers, we genotyped 60 Rhizophora racemosa drift propagules obtained on a bare unforested coastal area in southern Cameroon, estimated their relationship to 109 adult trees from most proximate sites (which were 3–85 km away), and assessed their relative difference with 873 trees of major mangrove areas (> 300 km) along the Cameroonian coastline. Proximate mangrove populations were considered as potential source populations in assignment tests. However, drift propagules could not be assigned to any of the Cameroonian mangrove sites and were genetically isolated from Cameroonian populations. Drift propagules showed higher levels of genetic diversity and private alleles giving a higher relatedness to each other than to any putative source population. Chloroplast sequences were used to confirm the identity of drift propagules as R. racemosa. We postulate that a complex interaction of ocean currents, estuarine geomorphology, and tidal patterns explain drift propagule dispersal to an area. Most likely the investigated cohort of propagules originated from more southern mangrove areas of the West African range beyond the Cameroonian border. This study unraveled the allelic, genetic, and genotypic features of stranded propagules following a stochastic long-distance dispersal. Transboundary dispersal of these propagules highlights the need for intergovernmental efforts in the management of biodiversity.

Development and evaluation of a custom bait design based on 469 single-copy protein-coding genes for exon capture of isopods (Philosciidae: Haloniscus).

Transcriptome-based exon capture approaches, along with next-generation sequencing, are allowing for the rapid and cost-effective production of extensive and informative phylogenomic datasets from non-model organisms for phylogenetics and population genetics research. These approaches generally employ a reference genome to infer the intron-exon structure of targeted loci and preferentially select longer exons. However, in the absence of an existing and well-annotated genome, we applied this exon capture method directly, without initially identifying intron-exon boundaries for bait design, to a group of highly diverse Haloniscus (Philosciidae), paraplatyarthrid and armadillid isopods, and examined the performance of our methods and bait design for phylogenetic inference. Here, we identified an isopod-specific set of single-copy protein-coding loci, and a custom bait design to capture targeted regions from 469 genes, and analysed the resulting sequence data with a mapping approach and newly-created post-processing scripts. We effectively recovered a large and informative dataset comprising both short (<100 bp) and longer (>300 bp) exons, with high uniformity in sequencing depth. We were also able to successfully capture exon data from up to 16-year-old museum specimens along with more distantly related outgroup taxa, and efficiently pool multiple samples prior to capture. Our well-resolved phylogenies highlight the overall utility of this methodological approach and custom bait design, which offer enormous potential for application to future isopod, as well as broader crustacean, molecular studies.

SNP Development in Penaeus vannamei via Next-Generation Sequencing and DNA Pool Sequencing

Next-generation sequencing and pool sequencing have been widely used in SNP (single-nucleotide polymorphism) detection and population genetics research; however, there are few reports on SNPs related to the growth of Penaeus vannamei. The purpose of this study was to call SNPs from rapid-growing (RG) and slow-growing (SG) individuals’ transcriptomes and use DNA pool sequencing to assess the reliability of SNPs. Two parameters were applied to detect SNPs. One parameter was the p-values generated using Fisher’s exact test, which were used to calculate the significance of allele frequency differences between RG and SG. The other one was the AFI (minor allele frequency imbalance), which was defined to highlight the fold changes in MAF (minor allele frequency) values between RG and SG. There were 216,015 hypothetical SNPs, which were obtained based on the transcriptome data. Finally, 104 high-quality SNPs and 96,819 low-quality SNPs were predicted. Then, 18 high-quality SNPs and 17 low-quality SNPs were selected to assess the reliability of the detection process. Here, 72.22% (13/18) accuracy was achieved for high-quality SNPs, while only 52.94% (9/17) accuracy was achieved for low-quality SNPs. These SNPs enrich the data for population genetics studies of P. vannamei and may play a role in the development of SNP markers for future breeding studies.

De Novo Transcriptomic Characterization Enables Novel Microsatellite Identification and Marker Development in Betta splendens.

The wild populations of the commercially valuable ornamental fish species, Betta splendens, and its germplasm resources have long been threatened by habitat degradation and contamination with artificially bred fish. Because of the lack of effective marker resources, population genetics research projects are severely hampered. To generate genetic data for developing polymorphic simple sequence repeat (SSR) markers and identifying functional genes, transcriptomic analysis was performed. Illumina paired-end sequencing yielded 105,505,486 clean reads, which were then de novo assembled into 69,836 unigenes. Of these, 35,751 were annotated in the non-redundant, EuKaryotic Orthologous Group, Swiss-Prot, Kyoto Encyclopedia of Genes and Genomes and Gene Ontology databases. A total of 12,751 SSR loci were identified from the transcripts and 7970 primer pairs were designed. One hundred primer pairs were randomly selected for PCR validation and 53 successfully generated target amplification products. Further validation demonstrated that 36% (n = 19) of the 53 amplified loci were polymorphic. These data could not only enrich the genetic information for the identification of functional genes but also effectively facilitate the development of SSR markers. Such knowledge would accelerate further studies on the genetic variation and evolution, comparative genomics, linkage mapping and molecular breeding in B. splendens.

"Jumping Jack": Genomic Microsatellites Underscore the Distinctiveness of Closely Related Pseudoperonospora cubensis and Pseudoperonospora humuli and Provide New Insights Into Their Evolutionary Past.

Downy mildews caused by obligate biotrophic oomycetes result in severe crop losses worldwide. Among these pathogens, Pseudoperonospora cubensis and P. humuli, two closely related oomycetes, adversely affect cucurbits and hop, respectively. Discordant hypotheses concerning their taxonomic relationships have been proposed based on host–pathogen interactions and specificity evidence and gene sequences of a few individuals, but population genetics evidence supporting these scenarios is missing. Furthermore, nuclear and mitochondrial regions of both pathogens have been analyzed using microsatellites and phylogenetically informative molecular markers, but extensive comparative population genetics research has not been done. Here, we genotyped 138 current and historical herbarium specimens of those two taxa using microsatellites (SSRs). Our goals were to assess genetic diversity and spatial distribution, to infer the evolutionary history of P. cubensis and P. humuli, and to visualize genome-scale organizational relationship between both pathogens. High genetic diversity, modest gene flow, and presence of population structure, particularly in P. cubensis, were observed. When tested for cross-amplification, 20 out of 27 P. cubensis-derived gSSRs cross-amplified DNA of P. humuli individuals, but few amplified DNA of downy mildew pathogens from related genera. Collectively, our analyses provided a definite argument for the hypothesis that both pathogens are distinct species, and suggested further speciation in the P. cubensis complex.

HLA diversity in the Russian population assessed by next generation sequencing

Next generation sequencing is used to determine full-length sequences of HLA genes at the 4-field (allelic) resolution. The study was aimed at determining frequency and diversity of HLA alleles in a cohort of blood donors from the Registry of the National Research Center for Hematology who design ated themselves as Russians (including some not routinely typed variations in HLA gene regions). The studied population consisted of 1510 donors. HLA typing was performed by next generation sequencing. Libraries were performed with AllType NGS Amplification Kits (One Lambda, USA) and sequenced using MiSeq (Illumina, USA). Data analysis used the TypeStream Visual Software V2.0.0.68 (One Lambda, USA) and IPD-IMGT/HLA database 3.40.0.1. Arlequin 3.5 software was used for estimation of allele and haplotype frequencies, deviation from Hardy-Weinberg equilibrium. 82 HLA-A, 156 HLA-В and 85 HLA-С alleles were identified with four-field resolution. 45 HLA-DRB1 and 18 HLA-DQB1 alleles were identified with 2-3-field resolution. Considerable HLA diversity was found among the donors self-designated as Russians: the population had large numbers of distinct alleles at each HLA gene, high percentage of alleles (25-32% of HLA class I) were revealed only once. Sufficient numbers of new alleles were registered which are absent in the IPD-IMGT/HLA database. Considerable allelic diversity in Russian population is due to low-incidence alleles. Despite this diversity, the majority of HLA alleles detected at each locus were common. Significant HLA diversity of the donors was connected with a large number of alleles with rare occurrence. The novel alleles identified in our study differed from the known alleles by single nucleotide substitutions. The most common alleles at the four-field level were as follows: A*02:01:01:01 (27.1%), C*07:02:01:03 (13.1%), A*03:01:01:01 (13.0%), B*07:02:01:01 (13.0%), A*01:01:01:01 (11.6%) and C*07:01:01:01/16 (10.4%). The HLA alleles, which are common for Russian populations, are not always common or well-documented alleles in present catalogues. The data obtained in this study may be used as a reference sample for estimation of HLA allele frequencies in Russian population, for proper frequency evaluation of specific alleles when searching donors for allogeneic hematopoietic stem cell transplantation, as well as for association studies between HLA alleles and different diseases, and for research in population genetics.

Assessing genotyping errors in mammalian museum study skins using high-throughput genotyping-by-sequencing

The use of museum specimens held in natural history repositories for population and conservation genetic research is increasing in tandem with the use of massively parallel sequencing technologies. Short Tandem Repeats (STRs), or microsatellite loci, are commonly used genetic markers in wildlife and population genetic studies. However, they traditionally suffered from a host of issues including length homoplasy, high costs, low throughput, and difficulties in reproducibility across laboratories. Massively parallel sequencing technologies can address these problems, but the incorporation of museum specimen derived DNA suffers from significant fragmentation and exogenous DNA contamination. Combatting these issues requires extra measures of stringency in the lab and during data analysis, yet there have not been any high-throughput sequencing studies evaluating microsatellite allelic dropout from museum specimen extracted DNA. In this study, we evaluate genotyping errors derived from mammalian museum skin DNA extracts for previously characterized microsatellites across PCR replicates utilizing high-throughput sequencing. We found it useful to classify samples based on DNA concentration, which determined the rate by which genotypes were accurately recovered. Longer microsatellites performed worse in all museum specimens. Allelic dropout rates across loci were dependent on sample quantity, with high concentration museum specimens performing as well and recovering quality metrics nearly as high as the frozen tissue sample. Based on our results, we provide a set of best practices for quality assurance and incorporation of reliable genotypes from museum specimens.

Genetic polymorphism of 23 autosomal STR loci in Han population from Yuncheng, Shanxi Province.

Due to the genetic feature of high diversity than other DNA markers, short tandem repeat (STR) plays key roles in forensic, anthropology, and population genetics. Newly introduced multiple STR kit is more valuable because of the greatly improved discriminatory power with the increase in the number of STR loci. The genetic polymorphic data are essential for the application and research in specific population. This study aims to investigate the genetic polymorphism of Han population residing in Yuncheng district, Shanxi Province, to evaluate the application of 23 STR loci in forensic personal identification and paternity test, and to explore the genetic relationship of Han population between Yuncheng and other populations. A total of 23 STR loci were amplified from 525 healthy unrelated individuals from the Han nationality in Yuncheng, Shanxi Province using the AGCU EX25 amplification kit. The products were detected and separated by ABI 3500 Genetic Analyzer. Alleles were genotyped by GeneMapper ID (Version 3.2) software, and corresponding frequencies and forensic parameters were calculated. We calculated the genetic distance and plotted the neighboring-joining tree with other 13 population. The allele frequency of the 23 STRs ranged from 0.0010 to 0.5090. No deviation from Hardy-Weinberg equilibrium (P>0.05) and linkage disequilibrium was observed. The cumulative discriminatory power (CPD), cumulative power of exclusion for trios (CPEtrio)and cumulative Power of exclusion for duos (CPEduo) with total 23 STRs were 1-1.305 263 374 8×10-27, 1-2.583 152 052 2×10-10 and 1-1.193 637 500 4×10-6, respectively. Comprehensive population comparison showed that Shanxi Yuncheng Han nationality was genetically closer to populations of the same linguistic family or geographically close proximity, such as Shaanxi Weinan Han, Liaoning Han, and Ningbo Han nationality while relatively far away from different linguistic ethnic groups and geographically distant populations like Xinjiang Uygur and Guangdong Han nationality. These 23 STRs are highly genetic polymorphic and informative in the Han population of Yuncheng, Shanxi Province, which can provide basic data for forensic personal identification, paternity testing, and population genetic research.

Genetic diversity analysis of Chinese plum (Prunus salicina L.) based on whole-genome resequencing

Chinese plum (Prunus salicina L.), also known as Japanese plum, is gaining importance because of its extensive genetic diversity and nutritional attributes that are beneficial for human health. Single-nucleotide polymorphisms (SNPs) are the most abundant form of genomic polymorphisms and are widely used in population genetics research. In this study, we constructed high-quality SNPs through whole-genome resequencing of 67 Prunus accessions with a depth of ~20× to evaluate the genome-level diversity and population structure. Phylogenetic analysis, principal component analysis, and population structure profiling indicated that the 67 plum accessions could be classified into four groups corresponding to their origin location, the southern cultivar group (SCG), the northern cultivar group (NCG), the foreign cultivar group (FG), and the mixed cultivar group (MG). Some cultivars from South China clustered with the other three groups. The genetic diversity indices including private allele number, observed heterozygosity, expected heterozygosity, and the nucleotide diversity of the SCG were higher than those of the NCG. Gene flow from the SCG to FG was also detected. Based on the distribution of wild resources, we concluded that the domestication center of origin of the Chinese plum was southwestern China. This study also provided genetic variation features and the population structure of Chinese plum cultivars, laying a foundation for breeders to use diverse germplasm and allelic variants to improve Chinese plum varieties.

PopAmaranth: a population genetic genome browser for grain amaranths and their wild relatives.

The combination of genomic, physiological, and population genetic research has accelerated the understanding and improvement of numerous crops. For nonmodel crops, the lack of interdisciplinary research hinders their improvement. Grain amaranth is an ancient nutritious pseudocereal that has been domesticated three times in different regions of the Americas. We present and employ PopAmaranth, a population genetic genome browser, which provides an accessible representation of the genetic variation of the three-grain amaranth species (Amaranthus hypochondriacus, Amaranthus cruentus, and Amaranthus caudatus) and two wild relatives (Amaranthus hybridus and Amaranthus quitensis) along the A. hypochondriacus reference sequence. We performed population-scale diversity and selection analysis from whole-genome sequencing data of 88 curated genetically and taxonomically unambiguously classified accessions. We employ the platform to show that genetic diversity in the water stress-related MIF1 gene declined during amaranth domestication and provide evidence for convergent saponin reduction between amaranth and quinoa. PopAmaranth is available through amaranthGDB at amaranthgdb.org/popamaranth.html

Bioinformatics analysis of microsatellite sites in the RNA-sequencing of &amp;lt;italic&amp;gt;Meretrix meretrix&amp;lt;/italic&amp;gt;

<italic>Meretrix meretrix</italic> is one of the most important economic shellfish species in China due to rich nutrition, delicious meat and high economic value. In this study, <italic>Meretrix meretrix</italic> DNA was transcribed and sequenced by using the second generation high-throughput sequencing technique. Through the statistical analysis of different types of microsatellites in <italic>Meretrix meretrix</italic>, the distribution frequency and number of microsatellites were understood, and the SSR composition characteristics of <italic>Meretrix meretrix</italic> were clarified at the transcriptome level. It is beneficial to the research of individual identification, germplasm resource identification, biological genetic mapping and so on, and has important reference value for further using microsatellite marker technology to carry out genetic marker-assisted breeding and population genetics research. The bioinformatics method was used to analyze the distribution of microsatellite sequences in the transcriptome of <italic>Meretrix meretrix</italic>. After searching the SSR loci of <italic>Meretrix meretrix</italic>, 8 856 complete SSR loci were found. The total length of SSR was 162 416 bp, with an average length of 18.34 bp, accounting for about 0.15% of the total sequence size, including 3 001 single-base repeat sequences, 2 254 three-base repeat sequences, 2 200 four-base repeats sequence, 1 052 two-base repeats sequences, 332 five-base repeat sequences and 13 six-base repeat sequences. The SSR of <italic>Meretrix meretrix</italic> transcriptome contained 26 types of repeats, among which the most was single-base repeat sequence A / T (2 809). The second most was triple-base repeat sequence AAC / TTG (907). Four-base repeat sequence AAAC / TTTG and two-base repeat sequence AT / TA was 588 and 561, respectively, indicating that the distribution of microsatellites in the RNA-sequencing of <italic>Meretrix meretrix</italic> had a preference for A / T. The average length of SSR (integrity) of <italic>Meretrix meretrix</italic> was 18.34 bp, and the main length of microsatellite was 12-20 bp, accounting for 41%. Most of the 8 856 SSR loci in the transcriptome had moderate polymorphism potential, which meant certain value for the future development of microsatellite molecular markers. The results of this study will provide a theoretical basis for the development of microsatellite markers, population genetic diversity, genetic linkage map, identification of germplasm resources and molecular genetics and breeding of shellfish. It also has important reference value for further study of genetic marker-assisted breeding and population genetics by using microsatellite marker technology.

Double-digest RAD-sequencing: do pre- and post-sequencing protocol parameters impact biological results?

Next-generation sequencing technologies have opened a new era of research in population genetics. Following these new sequencing opportunities, the use of restriction enzyme-based genotyping techniques, such as restriction site-associated DNA sequencing (RAD-seq) or double-digest RAD-sequencing (ddRAD-seq), has dramatically increased in the last decade. From DNA sampling to SNP calling, the laboratory and bioinformatic parameters of enzyme-based techniques have been investigated in the literature. However, the impact of those parameters on downstream analyses and biological results remains less documented. In this study, we investigated the effects of sevral pre- and post-sequencing settings on ddRAD-seq results for two biological systems: a complex of butterfly species (Coenonympha sp.) and several populations of common beech (Fagus sylvatica). Our results suggest that pre-sequencing parameters (i.e., DNA quantity, number of PCR cycles during library preparation) have a significant impact on the number of recovered reads and SNPs, on the number of unique alleles and on individual heterozygosity. In the same way, we found that post-sequencing settings (i.e., clustering and minimum coverage thresholds) influenced loci reconstruction (e.g., number of loci, mean coverage) and SNP calling (e.g., number of SNPs; heterozygosity) but had only a marginal impact on downstream analyses (e.g., measure of genetic differentiation, estimation of individual admixture, and demographic inferences). In addition, replication analyses confirmed the reproducibility of the ddRAD-seq procedure. Overall, this study assesses the degree of sensitivity of ddRAD-seq data to pre- and post-sequencing protocols, and illustrates its robustness when studying population genetics.

A likelihood ratio approach for identifying three-quarter siblings in genetic databases

The detection of family relationships in genetic databases is of interest in various scientific disciplines such as genetic epidemiology, population and conservation genetics, forensic science, and genealogical research. Nowadays, screening genetic databases for related individuals forms an important aspect of standard quality control procedures. Relatedness research is usually based on an allele sharing analysis of identity by state (IBS) or identity by descent (IBD) alleles. Existing IBS/IBD methods mainly aim to identify first-degree relationships (parent–offspring or full siblings) and second degree (half-siblings, avuncular, or grandparent–grandchild) pairs. Little attention has been paid to the detection of in-between first and second-degree relationships such as three-quarter siblings (3/4S) who share fewer alleles than first-degree relationships but more alleles than second-degree relationships. With the progressively increasing sample sizes used in genetic research, it becomes more likely that such relationships are present in the database under study. In this paper, we extend existing likelihood ratio (LR) methodology to accurately infer the existence of 3/4S, distinguishing them from full siblings and second-degree relatives. We use bootstrap confidence intervals to express uncertainty in the LRs. Our proposal accounts for linkage disequilibrium (LD) by using marker pruning, and we validate our methodology with a pedigree-based simulation study accounting for both LD and recombination. An empirical genome-wide array data set from the GCAT Genomes for Life cohort project is used to illustrate the method.

Efficient genotyping with backwards compatibility: converting a legacy microsatellite panel for muskellunge (Esox masquinongy) to genotyping-by-sequencing chemistry

Microsatellites have been a staple of population genetics research for over three decades, and many large datasets have been generated with these markers. Microsatellites have been used, for example, to conduct genetic monitoring and construct large multigeneration pedigrees as well as genotype thousands of individuals from a given species to create high-resolution baselines of spatial genetic structure. However, the capillary electrophoresis (CE) approach used to genotype microsatellites is inefficient compared to newer genotyping-by-sequencing (GBS) approaches, and researchers have begun transitioning away from CE. Backward compatibility between GBS and CE would facilitate a seamless transition to a more efficient chemistry, while ensuring that research based on CE panels could continue. Here, we explore the feasibility of converting a legacy panel of 15 microsatellites developed for muskellunge (Esox masquinongy) from CE to GBS chemistry. Muskellunge are an important sportfish in the Great Lakes region, and the existing microsatellite panel has been used to genotype thousands of samples to develop a region-wide baseline of genetic structure. We successfully converted all 15 microsatellites to GBS chemistry. GBS produced high genotyping rates (98%) and had high concordance with CE microsatellite genotypes (99%). Conversion to GBS required redesign of some primers and pairs to shorten amplicon length and adjust melting temperatures, optimization of primer concentrations, and comparisons with CE genotypes to optimize GBS genotyping parameters; however, none of these steps were especially onerous. Our results demonstrate that it is highly feasible to convert legacy CE panels to GBS, ensuring seamless continuation of important, often long-term research.

A duplex probe-directed recombinase amplification assay for detection of single nucleotide polymorphisms on 8q24 associated with prostate cancer.

Single nucleotide polymorphisms (SNPs) have important application value in the research of population genetics, hereditary diseases, tumors, and drug development. Conventional methods for detecting SNPs are typically based on PCR or DNA sequencing, which is time-consuming, costly, and requires complex instrumentation. In this study, we present a duplex probe-directed recombinase amplification (duplex-PDRA) assay that can perform real-time detection of two SNPs (rs6983267 and rs1447295) in four reactions in two tubes at 39°C within 30 min. The sensitivity of duplex-PDRA was 2×103-104 copies per reaction and no cross-reactivity was observed. A total of 382 clinical samples (179 prostate cancer patients and 203 controls) from northern China were collected and tested by duplex-PDRA assay and direct sequencing. The genotyping results were completely identical. In addition, the association analysis of two SNPs with prostate cancer risk and bone metastasis was conducted. We found that the TT genotype of rs6983267 (OR: 0.42; 95%CI: 0.23-0.78; P=0.005) decreased the risk of prostate cancer, while the CA genotype of rs1447295 (OR: 1.89; 95%CI: 1.20-2.96; P=0.005) increased the risk of prostate cancer. However, no association between the two SNPs (rs6983267 and rs1447295) and bone metastasis in prostate cancer was found in this study (P>0.05). In conclusion, the duplex-PDRA assay is an effective method for the simultaneous detection of two SNPs and shows great potential for widespread use in research and clinical settings.

Comparing Genetic Variation among Latin American Immigrants: Implications for Forensic Casework in the Arizona- and Texas-Mexico Borderlands.

The humanitarian crisis on the US-Mexico border is a long-standing and evolving crisis in which nearly 8,000 deaths have been reported in the last two decades. These deaths are largely distributed across the Arizona-Mexico and Texas-Mexico border regions, where demographic trends for immigrants attempting to cross into the United States have shifted dramatically. The demographic change and volume of immigrants seeking shelter in the United States present new challenges for the forensic practitioners entrusted with the identification of individuals who lose their lives during the final segment of their journey. Within this border context, this study investigated how genetic variation inferred from forensically significant microsatellites can provide valuable information on regions of origin for unidentified remains at the group level. To explore how to mobilize these genetic data to inform identification strategies, the authors conducted a comparative genetic analysis of identified and unidentified immigrant cases from the Arizona- and Texas-Mexico contexts, as well as 27 other Latin American groups. Allele frequencies were utilized to calculate FST, and relationships were visually depicted in a multidimensional scaling plot. A Spearman correlation coefficient analysis assessed the strength and significance of population relationships, and an agglomerative clustering analysis assessed population clusters. Results indicate that Arizona-Mexico immigrants have the strongest relationship (>80%) with groups from El Salvador, Guatemala, Mexico, and an indigenous group from southern Mexico. Texas-Mexico immigrants have the strongest relationships (>80%) with groups from Belize, Colombia, Costa Rica, El Salvador, Guatemala, Honduras, and Nicaragua. These findings agree with, and are discussed in comparison with, previously reported demographic trends, population genetics research, and population history analyses. The authors emphasize the utility and necessity of coupling genetic variation research with a nuanced anthropological perspective for identification processes in the US-Mexico border context.