Kinship Analysis Research Articles

DNA-based genealogy reconstruction of Nebbiolo, Barbera and other ancient grapevine cultivars from northwestern Italy

Northwestern Italy is a wine region of the world with the highest of reputations, where top quality wines of remarkable economic value are produced from traditional, long-cultivated varieties. Kinship analyses were performed using 32 microsatellite loci and more than 10 K single-nucleotide polymorphism markers on 227 traditional grapes mostly from Northwestern Italy—including those that have been neglected or are threatened. This was done to better understand the genetic grapevine origins and history of this reputable wine producing area, thus enhancing its cultural value and the marketing appeal of its wines. The work revealed a complex network of genetic relationships among varieties, with little contribution of genotypes from other areas. It revealed the major role played by a few ancient grape varieties as parents of numerous offspring, including some that are endangered today. The ancestry of many cultivars is proposed. Among these are Dolcetto, Barbera and Riesling italico. Through the inference of parent–offspring and sibling relations, marker profiles of ungenotyped putative parents were reconstructed, suggesting kinship relations and a possible parentage for Nebbiolo, one of the most ancient wine grapes worldwide. Historic and geographic implications from the resulting kinships are discussed.

Autosomal STR Profiling and Databanking in Malaysia: Current Status and Future Prospects.

Science and technology are extensively used in criminal investigation. From the mid- to late-1980s, one of the scientific discoveries that has had a particularly remarkable impact on this field has been the use of highly variable DNA sequence regions (minisatellites) in the human genome for individual identification. The technique was initially referred to as DNA fingerprinting, but is now more widely referred to as DNA profiling. Since then, many new developments have occurred within this area of science. These include the introduction of new genetic markers (microsatellites also known as short tandem repeats/STRs), the use of the polymerase chain reaction for target amplification, the development of DNA databases (databanking), and the advancement and/or improvement of genotyping protocols and technologies. In 2019, we described the progress of DNA profiling and DNA databanking in Malaysia for the first time. This report included information on DNA analysis regulations and legislation, STR genotyping protocols, database management, and accreditation status. Here, we provide an update on the performance of our DNA databank (numbers of DNA profiles and hits) plus the technical issues associated with correctly assigning the weight of evidence for DNA profiles in an ethnically diverse population, and the potential application of rapid DNA testing in the country. A total of 116,534 DNA profiles were obtained and stored in the Forensic DNA Databank of Malaysia (FDDM) by 2019, having increased from 70,570 in 2017. The number of hits increased by more than three-fold in just two years, where 17 and 69 hits between the DNA profiles stored in the FDDM and those from crime scenes, suspects, detainees, drug users, convicts, missing persons, or volunteers were recorded in 2017 and 2019, respectively. Forensic DNA analysis and databanking are thus progressing well in Malaysia and have already contributed to many criminal investigations. However, several other issues are discussed here, including the need for STR population data for uncharacterized population groups, and pilot trials for adopting rapid DNA profiling technology. These aspects should be considered by policy makers and law enforcement agencies in order to increase the reliability and efficiency of DNA profiling in criminal cases and in kinship analysis in Malaysia.

Resequencing of 1,143 indica rice accessions reveals important genetic variations and different heterosis patterns

Obtaining genetic variation information from indica rice hybrid parents and identification of loci associated with heterosis are important for hybrid rice breeding. Here, we resequence 1,143 indica accessions mostly selected from the parents of superior hybrid rice cultivars of China, identify genetic variations, and perform kinship analysis. We find different hybrid rice crossing patterns between 3- and 2-line superior hybrid lines. By calculating frequencies of parental variation differences (FPVDs), a more direct approach for studying rice heterosis, we identify loci that are linked to heterosis, which include 98 in superior 3-line hybrids and 36 in superior 2-line hybrids. As a proof of concept, we find two accessions harboring a deletion in OsNramp5, a previously reported gene functioning in cadmium absorption, which can be used to mitigate rice grain cadmium levels through hybrid breeding. Resource of indica rice genetic variation reported in this study will be valuable to geneticists and breeders.

Kinship analysis on single cells after whole genome amplification

Short Tandem Repeat (STR-) and Single Nucleotide Polymorphism (SNP-) genotyping have been extensively studied within forensic kinship analysis. Nevertheless, no results have been reported on kinship analysis after whole genome amplification (WGA) of single cells. This WGA step is a necessary procedure in several applications, such as cell-based non-invasive prenatal testing (cbNIPT) and pre-implantation genetic diagnosis (PGD). In cbNIPT, all putative fetal cells must be discriminated from maternal cells after enrichment from whole blood. This study investigates the efficacy and evidential value of STR- and SNP-genotyping methods for the discrimination of 24 single cells after WGA, within three families. Formaldehyde-fixed and unfixed cells are assessed in offspring-parent duos and offspring-mother-father trios. Results demonstrate that both genotyping methods can be used in all tested conditions and scenarios with 100% sensitivity and 100% specificity, with a similar evidential value for fixed and unfixed cells. Moreover, sequence-based SNP-genotyping results in a higher evidential value than length-based STR-genotyping after WGA, which is not observed using high-quality offspring bulk DNA samples. Finally, it is also demonstrated that the availability of the DNA genotypes of both parents strongly increases the evidential value of the results.

A Forensic Genomics Approach for the Identification of Sister Marija Crucifiksa Kozulić.

Sister Marija Krucifiksa Kozulić (1852–1922) was a Croatian nun who is in consideration for beatification by the Vatican, which is facilitated by the identification of her 20th-century remains. Sister Marija was buried in a tomb in Rijeka, Croatia, along with other nuns including her biological sister, Tereza Kozulić (1861–1933). When the remains were exhumed in 2011, they were found in a deteriorated state and commingled with several other sets of remains. Thus, mitochondrial genome sequencing of the long bones was performed to sort the remains by mitochondrial haplotype. Two similar but unique haplotypes belonging to haplogroup H1bu were identified, and samples from these bones were subjected to autosomal short tandem repeat (STR) and single nucleotide polymorphism (SNP) sequencing. Although only partial profiles were obtained, the data were sufficient for kinship analysis with the profile of a paternal niece of Sister Marija (Fides Kozulić). The data indicate that it is 574,195-fold more likely that the two sets of skeletal remains represent 2nd-degree relatives of Fides than sisters who are unrelated to Fides. Although it is impossible to discern which set of remains belongs to Marija and which belongs to Tereza, forensic genomics methods have enabled identification of the sisters.

Genetic evidence suggests a sense of family, parity and conquest in the Xiongnu Iron Age nomads of Mongolia.

In an effort to characterize the people who composed the groups known as the Xiongnu, nuclear and whole mitochondrial DNA data were generated from the skeletal remains of 52 individuals excavated from the Tamir Ulaan Khoshuu (TUK) cemetery in Central Mongolia. This burial site, attributed to the Xiongnu period, was used from the first century BC to the first century AD. Kinship analyses were conducted using autosomal and Y-chromosomal DNA markers along with complete sequences of the mitochondrial genome. These analyses suggested close kin relationships between many individuals. Nineteen such individuals composed a large family spanning five generations. Within this family, we determined that a woman was of especially high status; this is a novel insight into the structure and hierarchy of societies from the Xiongnu period. Moreover, our findings confirmed that the Xiongnu had a strongly admixed mitochondrial and Y-chromosome gene pools and revealed a significant western component in the Xiongnu group studied. Using a fine-scale approach (haplotype instead of haplogroup-level information), we propose Scytho-Siberians as ancestors of the Xiongnu and Huns as their descendants.

Travel with your kin ship! Insights from genetic sibship among settlers of a coral damselfish.

Coral reef fish larvae are tiny, exceedingly numerous, and hard to track. They are also highly capable, equipped with swimming and sensory abilities that may influence their dispersal trajectories. Despite the importance of larval input to the dynamics of a population, we remain reliant on indirect insights to the processes influencing larval behavior and transport. Here, we used genetic data (300 independent single nucleotide polymorphisms) derived from a light trap sample of a single recruitment event of Dascyllus abudafur in the Red Sea (N = 168 settlers). We analyzed the genetic composition of the larvae and assessed whether kinship among these was significantly different from random as evidence for cohesive dispersal during the larval phase. We used Monte Carlo simulations of similar‐sized recruitment cohorts to compare the expected kinship composition relative to our empirical data. The high number of siblings within the empirical cohort strongly suggests cohesive dispersal among larvae. This work highlights the utility of kinship analysis as a means of inferring dynamics during the pelagic larval phase.

One panel to rule them all: DArTcap genotyping for population structure, historical demography, and kinship analyses, and its application to a threatened shark.

With recent advances in sequencing technology, genomic data are changing how important conservation management decisions are made. Applications such as Close-Kin Mark-Recapture demand large amounts of data to estimate population size and structure, and their full potential can only be realised through ongoing improvements in genotyping strategies. Here we introduce DArTcap, a cost-efficient method that combines DArTseq and sequence capture, and illustrate its use in a high resolution population analysis of Glyphis garricki, a rare, poorly known and threatened euryhaline shark. Clustering analyses and spatial distribution of kin pairs from four different regions across northern Australia and one in Papua New Guinea, representing its entire known range, revealed that each region hosts at least one distinct population. Further structuring is likely within Van Diemen Gulf, the region that included the most rivers sampled, suggesting additional population structuring would be found if other rivers were sampled. Coalescent analyses and spatially explicit modelling suggest that G.garricki experienced a recent range expansion during the opening of the Gulf of Carpentaria following the conclusion of the Last Glacial Maximum. The low migration rates between neighbouring populations of a species that is found only in restricted coastal and riverine habitats show the importance of managing each population separately, including careful monitoring of local and remote anthropogenic activities that may affect their environments. Overall we demonstrated how a carefully chosen SNP panel combined with DArTcap can provide highly accurate kinship inference and also support population structure and historical demography analyses, therefore maximising cost-effectiveness.

ANALISIS KERAGAMAN DAN KEKERABATAN KACANG TUNGGAK (Vigna unguiculata L) GENERASI M2

Cowpea (V. unguiculata L) has great potential as a nutritious food as a substitute for soybeans because it contains sufficient protein and low fat content. The diversity of cowpea is low so that need to increase diversity through the mutation induction of gamma ray irradiation. The study was carried out in the experimental garden of Pasir Kuda PKHT of IPB in February - May 2018 using a design of augmented in the Complete Group Design in a Randomized. The study was carried out using 90 putative mutant genotypes of M2 generation as the test genotype and KM4 genotype as a comparison which was repeated 10 times. The results showed that the M2 generation putative mutants showed diversity in the qualitative and quantitative characters of cowpea. High genetic diversity is shown in the character of plant height, harvest period, number of seeds / pods and weight of cowpea seeds / plants and high broad mean heritability values obtained on the character of stem length, flowering age, number of seeds / pods and weight of beans / plant nuts arrears. The result of kinship analysis showed thirteen different putative mutant genotypes with KM4 genotypes, namely T6599P, T8028P, T7525P, T7551P, T7520P, T6574P, T6533P, T7058P, T6577P, T6591P, T7062P, T7069P and T6561.

Genetic diversity of X-STR markers in Jat Sikh population of Punjab, India and its comparison with other 39 global populations

Background The Jat Sikh population is the largest endogamous group of Punjab, a state in north-west India, and has not yet been explored for genetic polymorphism based on X-STR genetic markers. In India, which is the second most populous country in the world, only two population studies based on X-STR markers have been reported so far. Aim To explore the genetic diversity of 12 X chromosomal STR genetic markers in the Jat Sikh population of Punjab and expand the X-STR polymorphism database. Subjects and methods In this study, a total of 200 Jat Sikh individuals (100 males and 100 females) residing in Punjab were investigated for 12 X-STR markers using the Investigator Argus X-12 QS Kit. Results The highest power of discrimination (PD) in females (PDf) and males (PDm) was observed to be 0.965 (DXS10135) and 0.929 (DXS10135 and DXS10148), respectively. DXS10135 was found to be the most polymorphic and discriminating locus among all the studied loci in both males and females with highest values of power of discrimination (PD) and polymorphic information content (PIC) as well. Conclusion Overall, the studied markers of the Argus 12 X-STR kit provide high polymorphic information which may prove to be an important tool in resolving issues such as missing person identification, incest, immigration disputes, kinship analysis and genealogical studies. The dataset obtained from this study will add to the present database of X-STRs.

Molecular genetic analysis of spring wheat core collection using genetic diversity, population structure, and linkage disequilibrium

BackgroundWheat (Triticum aestivium L.) is an important crop globally which has a complex genome. To identify the parents with useful agronomic characteristics that could be used in the various breeding programs, it is very important to understand the genetic diversity among global wheat genotypes. Also, understanding the genetic diversity is useful in breeding studies such as marker-assisted selection (MAS), genome-wide association studies (GWAS), and genomic selection.ResultsTo understand the genetic diversity in wheat, a set of 103 spring wheat genotypes which represented five different continents were used. These genotypes were genotyped using 36,720 genotyping-by-sequencing derived SNPs (GBS-SNPs) which were well distributed across wheat chromosomes. The tested 103-wheat genotypes contained three different subpopulations based on population structure, principle coordinate, and kinship analyses. A significant variation was found within and among the subpopulations based on the AMOVA. Subpopulation 1 was found to be the more diverse subpopulation based on the different allelic patterns (Na, Ne, I, h, and uh). No high linkage disequilibrium was found between the 36,720 SNPs. However, based on the genomic level, D genome was found to have the highest LD compared with the two other genomes A and B. The ratio between the number of significant LD/number of non-significant LD suggested that chromosomes 2D, 5A, and 7B are the highest LD chromosomes in their genomes with a value of 0.08, 0.07, and 0.05, respectively. Based on the LD decay, the D genome was found to be the lowest genome with the highest number of haplotype blocks on chromosome 2D.ConclusionThe recent study concluded that the 103-spring wheat genotypes and their GBS-SNP markers are very appropriate for GWAS studies and QTL-mapping. The core collection comprises three different subpopulations. Genotypes in subpopulation 1 are the most diverse genotypes and could be used in future breeding programs if they have desired traits. The distribution of LD hotspots across the genome was investigated which provides useful information on the genomic regions that includes interesting genes.

Interpreting Mixture Profiles: Comparison between Precision ID GlobalFiler™ NGS STR Panel v2 and Traditional Methods.

Forensic investigation for the identification of offenders, recognition of human remains, and verification of family relationships requires the analysis of particular types of highly informative DNA markers, which have high discriminatory power and are efficient for typing degraded samples. These markers, called STRs (Short Tandem Repeats), can be amplified by multiplex-PCR (Polymerase Chain Reaction) allowing attainment of a unique profile through which it is possible to distinguish one individual from another with a high statistical significance. The rapid and progressive evolution of analytical techniques and the advent of Next-Generation Sequencing (NGS) have completely revolutionized the DNA sequencing approach. This technology, widely used today in the diagnostic field, has the advantage of being able to process several samples in parallel, producing a huge volume of data in a short time. At this time, although default parameters of interpretation software are available, there is no general agreement on the interpretation rules of forensic data produced via NGS technology. Here we report a pilot study aimed for a comparison between NGS (Precision ID GlobalFiler™ NGS STR Panel v2, Thermo Fisher Scientific, Waltham, MA, USA) and traditional methods in their ability to identify major and minor contributors in DNA mixtures from saliva and urine samples. A quantity of six mixed samples were prepared for both saliva and urine samples from donors. A total of 12 mixtures were obtained in the ratios of 1:2; 1:4; 1:6; 1:8; 1:10; and 1:20 between minor and major contributors. Although the number of analyzed mixtures is limited, our results confirm that NGS technology offers a huge range of additional information on samples, but cannot ensure a higher sensitivity in respect to traditional methods. Finally, the Precision ID GlobalFiler™ NGS STR Panel v2 is a powerful method for kinship analyses and typing reference samples, but its use in biological evidence should be carefully considered on the basis of the characteristics of the evidence.

Weight-of-evidence for DNA identification of missing persons and human remains using CODIS.

DNA Identification of unidentified human remains (UHR) is performed in Israel by comparing the UHR's short tandem repeat (STR) profiles to a national database of STR profiles taken from relatives of missing persons. Kinship analysis is performed using the CODIS 7.0 software and results are stated as a Joint Pedigree Likelihood Ratio (JPLR). The weight-of-evidence for JPLR has never been studied, making it difficult to interpret the meaning of specific values in terms of whether UHR are related to specific pedigrees. Therefore, the aim of this study was to statistically determine the practical meaning and context of the JPLR. We used 440 million pairs of simulated DNA profiles and 294 pairs of real ones from known siblings, parent/offspring and unrelated persons. A Score-Based Likelihood Ratio (SBLR) was empirically constructed, validated and compared to both JPLR and the LR produced by CODIS. Our results show that CODIS's JPLR and LR values for single-person pedigrees overestimate the level of support for both "parent/child" and "siblings" propositions relative to the "unrelated" proposition, by up to two orders of magnitude. A practical table is given for correcting this phenomenon, with statistical interpretation (i.e. SBLR) for each JPLR score, including verbal levels of propositional support ranging from "no support" (SBLR<2) to "extremely strong" (SBLR>1 Million).

Sequenced-based French population data from 169 unrelated individuals with Verogen’s ForenSeq DNA signature prep kit

Massively Parallel Sequencing (MPS) applied to forensic genetics allows the simultaneous analysis of hundreds of genetic markers and the access to full amplicon sequences which help to increase available allele diversity. Meanwhile, sequence variation within the repeat regions represents the majority of the allele diversity, flanking regions adjacent to the repeat core provide an additional degree of variation. The forensic genetics community needs access to population data, from relevant parts of the world that contain this new sequence diversity in order to perform statistical calculations. In this study, we report sequence-based Short Tandem Repeat (STR) and identity Single Nucleotide Polymorphism (iSNPs) allele data for 169 French individuals across 58 STRs and 92 SNPs included in the Verogen ForenSeq DNA Signature Prep kit. 42 STRs out of 58 showed an increased number of alleles due to sequence variation in the repeat motif and/or the flanking regions. D9S1122 showed the largest overall gain with an increase of observed heterozygosities of almost 25 %. The combined match probability combining 27 autosomal STRs and 91 identity SNPs was 1.11E-69. Sequence-based allele frequencies included in this publication will help forensic laboratories to increase the power of discrimination for identification, kinship analysis and mixture interpretation.

Haplotype variations in QTL for salt tolerance in Chinese wheat accessions identified by marker-based and pedigree-based kinship analyses

Most modern wheat cultivars were selected on the basis of yield-related indices measured under optimal fertilizer and irrigation inputs. With climate change, land degradation and salinity caused by sea water encroachment, wheat is increasingly subjected to environmental stress. Moreover, expanding urbanization increasingly encroaches upon prime agricultural land in countries like China, and alternative cropping areas must be found. Some of these areas have moderate constraining factors, such as salinity. Therefore, it is important to investigate whether current genetic materials and breeding procedures are maintaining adequate variability to address future problems caused by abiotic stress. In this study, a panel of 307 wheat accessions, including local landraces, exotic cultivars used in Chinese breeding programs and Chinese cultivars released during different periods since 1940, were subjected to a genome-wide association study to dissect the genetic basis of salinity tolerance. Both marker-based and pedigree-based kinship analyses revealed that favorable haplotypes were introduced in some exotic cultivars as well as a limited number of Chinese landraces from the 1940s. However, improvements in salinity tolerance during modern breeding are not as obvious as that of yield. To broaden genetic diversity for increasing salt tolerance, there is a need to refocus attention on local landraces that have high degrees of salinity tolerance and carry rare favorable alleles that have not been exploited in breeding.

German population data for 18 X-STRs: a hexaplex PCR adding two clusters of X-STRs to the Argus X-12 set and expanding the German haplotype databases.

In kinship analysis, large data sets with estimated haplotype frequencies for marker clusters are very important for the likelihood calculation. Practical use of the X-STRs demonstrated that in some complex kinship cases, the marker set of the Investigator Argus X-12 kit can be insufficient. This study aimed to extend the German data base of the Argus X-12 kit (1037 haplotypes) and for a cluster in Xq21 (806 haplotypes) with additional 700 male haplotypes and to include a further cluster in Xp22.3 to complete the X-STR marker set for complex kinship cases.

Diversidad genética de Oreomunnea mexicana (Juglandaceae), relicta del bosque de niebla de Sierra Juárez, Oaxaca. México

Oreomunnea mexicana es una especie relicta y amenazada, que constituye un elemento arbóreo dominante en el bosque de niebla de la Sierra Juárez, Oaxaca. La especie es emblemática y se encuentra incorporada en la cosmovisión de los habitantes de la región, donde, además, desempeña funciones ecológicas y servicios ecosistémicos muy importantes. El objetivo del presente trabajo fue analizar la diversidad y estructura genética, así como evaluar la estructura genética espacial a escala fina, utilizando marcadores moleculares con fines de conservación in situ. Se utilizaron marcadores dominantes denominados inter simple sequence repeat con los que se analizaron individuos de nueve poblaciones (25 por población) y 120 individuos de una parcela de 400 m2. Los resultados mostraron niveles altos de diversidad genética (He = 0.405 ± 0003) y moderados de flujo génico (Nm = 2.25) y baja diferenciación genética (P &lt; 0.001; 8%). La estructura genética espacial fina mostró una distribución aleatoria, mientras que el análisis de parentesco exhibió alta similitud genética entre los individuos analizados. Los niveles altos de diversidad sugieren que la especie no se encuentra degradada genéticamente, sin embargo, debido a su distribución fragmentada y alta vulnerabilidad ambiental, se sugiere para su conservación in situ, se dé prioridad para su protección inmediata a las poblaciones de El Relámpago (por su gran riqueza alélica), El Panteón y El Orquidiario debido a su baja diversidad genética y por la alta vulnerabilidad por perturbación antrópica y ambiental.

Development of an NGS panel containing 42 autosomal STR loci and the evaluation focusing on secondary kinship analysis.

High-throughput next-generation sequencing (NGS) is a feasible technique to detect considerably more markers and simultaneously obtain length and sequence information in a single reaction. In this study, we developed an NGS panel including 42 commonly used autosomal short tandem repeats (STRs) and amelogenin on the Illumina MiSeq FGx™. Sequencing accuracy was validated by the consistency of 2800M Control DNA detected using the ForenSeq™ DNA Signature Prep Kit and Sanger sequencing. Nomenclature incompatibility was found between NGS-STR and CE-STR typing at 9 loci (D3S3045, D6S477, D7S3048, D9S925, D14S608, D17S1290, D18S535, D21S1270, GATA198B05), despite the correct sequence. The difference was caused by the two different methods of identifying motif sequence and a one-to-one correspondence can be found. We evaluated the panel by investigating consistency, sequencing sensitivity and the effectiveness of the 2nd-degree relationship identification. Herein, we present sequencing results from 58 unrelated individuals of the Hebei Han population. The total discrimination power (TDP) and cumulative probability of exclusion for trio paternity testing (CPEtrio) of the 42 NGS-STR panels reached 1-2.84 × 10-57 and 1-9.87 × 10-21, respectively. By family simulation and likelihood ratio (LR) calculation, this panel was shown to have effectiveness for the 2nd-degree kinship identification similar to the ForenSeq™ DNA Signature Prep Kit and certain advantages compared with it due to the relatively small number of loci. As expected, it provides new data for the development of NGS-STR typing technology.

Large-scale identification of human bone remains via SNP microarray analysis with reference SNP database

Single nucleotide polymorphisms (SNPs) are valuable markers complementary to conventional forensic short tandem repeat (STR) markers in genetic typing, with potential advantages in challenging forensic casework. With the advent of high-throughput technologies, such as microarrays and massively parallel sequencing, the use of SNP typing has now expanded to large-scale forensic applications. Herein, a forensic case is presented to demonstrate the usefulness of SNP typing in identifying large-scale human bone remains with reference database construction. A total of 402 bone remains were recovered from an island in the Jeju Province of Korea where a massive disaster occurred in 1948. The first phase of the identification process was accomplished via conventional DNA typing methods including autosomal and Y-chromosomal STR typing, and mitochondrial DNA sequencing, which resulted in the identification of 74 of 402 remains. The second phase of the identification involved the remaining 327 unidentified remains using SNP typing as a supplementary tool based on Affymetrix resequencing array. The SNP markers of 782 family members were also analyzed and a reference database was constructed for comparison. An additional 51 bone remains were identified in the second phase. SNP data obtained from the supplementary genotyping yielded additional genetic information as well as contributed to kinship testing to determine the second degrees of relationship. In addition SNPs are useful in discriminating ambiguous relationship when only STR data are available. A software program developed for SNP typing system enabled efficient kinship analysis for large-scale forensic identification. The results and the casework are described and discussed.

Development of a software for kinship analysis considering linkage and mutation based on a Bayesian network

In forensic DNA testing, the number of tested short tandem repeat loci has increased owing to new multiplex kits with additional loci. Although this advancement provides improved discrimination power, the effects of linkage and mutation must be considered during kinship analysis. However, no software currently includes both of these effects. In this study, we developed new freeware called KinBN for kinship analysis based on a Bayesian network. The software is graphical-user-interface-based and calculates the likelihood ratios (LRs) at multiple loci considering the effects of linkage and mutation. In addition, the software can simulate the LR distribution according to the specified relationship. We confirmed the accuracy of KinBN by comparing its LRs with those of other software and evaluated the effects of linkage and mutation on the LRs. Our results indicate that KinBN is a useful tool for kinship analysis, particularly if expanded locus sets are used for DNA testing.