Amelogenin Gene Research Articles

Sexing of pre-implantation ovine embryos through polymerase chain reaction-based amplification of GAPDH, SRY and AMEL genes.

The ability to identify the sex of embryo and control of sex ratio has a great commercial importance to livestock industry. Prediction of embryonic sex could be useful in the management decisions of sex selection in breeding programs. Several methods have been attempted to determine the sex but the polymerase chain reaction (PCR)-based sexing method is generally favoured, as it is cost effective, simple and reliable. The aim of the present study was to identify sex of sheep embryos produced in vitro through amplification of glyceraldehyde 3-phosphate dehydrogenase (GAPDH), sex-determining region Y (SRY) and amelogenin genes present in genomic DNA (gDNA) of embryos through PCR. To avoid false interpretation of the result by no amplification of SRY in female embryos, a duplex PCR was approached to amplify combinedly SRY and GAPDH genes. Sex-specific blood was used in PCR as positive control. In vitro sheep embryos were produced as per standardized protocol of laboratory. Sexing of sex-specific blood and in vitro produced embryos were approached though PCR to amplify the respective genes using gDNA present in the sample without its traditional isolation. The accuracy of sex prediction for embryos was 100% by this procedure.

Naked eye Y amelogenin gene fragment detection using DNAzymes on a paper-based device

Nowadays, great efforts are made in developing new technology for rapid detection of specific DNA sequences, for environmental monitoring, forensic analysis and rapid biomedical diagnosis applications. Microfluidic paper-based analytical devices are suitable platforms for the development of point of care devices, due to their simple fabrication protocols, ease of use and low cost. Herein, a methodology for in situ detection of single strand DNA by using a colorimetric assay based on the formation of a DNAzyme within a paper substrate was developed. A DNAzyme that could only be formed in the presence of a specific sequence of the Y human amelogenin gene was designed. The performance of the DNAzyme was followed colorimetrically first in solution and then in paper substrates. The reaction was found to be specific to the Y fragment selected as analyte. The DNAzyme reaction on paper enabled the unequivocal colorimetric identification of the Y single strand DNA fragment both qualitatively, with the naked eye (143 ng), and quantitatively by image analysis (45.7 ng). As a proof of concept, a microfluidic paper-based device, pre-loaded with all DNAzyme reagents, was characterized and implemented for the simultaneous detection of X and Y single strand DNA fragments.

Alternatives to amelogenin markers for sex determination in humans and their forensic relevance.

Forensic DNA typing and subsequent molecular methods of sex determination in humans have been proven to be an imperious tool to criminal justice system. In current practice, most of the short tandem repeat (STR) based commercial kits contain amelogenin as the sexing marker. Amelogenin gene which contributes to the tooth enamel formation is present on both X and Y chromosome with a variation in base pair size. However, huge discrepancies have been observed with amelogenin based sex determination mostly due to X and Y deletion in the population and mutation in primer binding sites. Some ethnicities such as those in Indian population are affected badly with inappropriate sex determination by amelogenin marker due to the presence of high frequency of Y deletion in the population. Presence of PCR inhibitors, degradation in the DNA samples and presence of mixed DNA also contribute to the discrepancy in results obtained by amelogenin analysis. To overcome this problem, many alternative markers/techniques such as STS, SRY, TSPY, DXYS156, SNPs, DYZ1 and Next generation sequencing have been discussed in much detail with their respective pros and cons. In this regard, inclusion of one or more alternative markers along with amelogenin will decrease the anomalies in sex determination observed while using the amelogenin marker alone in forensic sample analysis.

Molecular characterization of mitochondrial Amerindian haplogroups and the amelogenin gene in human ancient DNA from three archaeological sites in Lambayeque - Peru.

Important pre-Inca civilizations, known by their great political and religious structures, inhabited the northern coast of Peru. Archeological and anthropological studies have shown that people from these villages have hierarchical strata, but the genetic structure has been poorly studied. Here, we aimed to perform a molecular characterization of the Amerindian maternal lineages and the amelogenin gene in skeletons collected from three archeological sites in Lambayeque. Ancient DNA (aDNA) samples were analyzed with conventional PCR to assess the nine-base pair (9 bp) deletion corresponding to mitochondrial haplogroup B and the identification of haplogroups A, C, and D were obtained with PCR-RFLP experiments. The sex was characterized via amplification of the AMEL(X/Y) locus. Haplogroup frequencies were compared with available data from other ancient and modern civilizations from the Peruvian coast and highlands using statistical methods. Our results showed that haplogroup C had the highest frequency, while haplogroup B showed variable diversity in the analyzed populations. The meta-analysis revealed a positive correlation among some coastal villages. We concluded that ancient populations analyzed in our study showed the presence of four Amerindian mitochondrial haplogroups, which is consistent with previous studies.

Determination of DNA of women and men in the blood meal of Aedes aegypti based on the amelogenin and SRY genes.

Determination of DNA of women and men in the blood meal of Aedes aegypti based on the amelogenin and SRY genes.

Погребальный памятник афанасьевской культуры Бертек-33 на плато Укок (Горный Алтай): результаты палеогенетического исследования

В статье представлены результаты палеогенетического исследования останков семи взрослых индивидов из погребального памятника афанасьевской культуры Бертек-33 (плато Укок, Республика Алтай, Россия) и их интерпретация с учетом археологического и палеоантропологического контекста. Проведен анализ четырех систем генетических маркеров: митохондриальной ДНК, полиморфного фрагмента гена амелогенина, аутосомных STR-локусов и STR-локусов Y-хромосомы. Полученные результаты свидетельствуют о доминировании в исследуемой группе населения западно-евразийских гаплогрупп митохондриальной ДНК (T, J, U5a, K, H) и однородности мужского генофонда, представленного вариантами гаплогруппы R1b Y-хромосомы. Данные по митохондриальной ДНК, Y-хромосоме, отдельным аутосомным маркерам указывают на западно-евразийский вектор генетических связей этой группы. Исследованная серия укладывается в рамки внутрипопуляционного разнообразия генофонда митохондриальной ДНК и Y-хромосомы афанасьевского населения Южной Сибири. В статье рассмотрены возможные родственные связи между индивидами, захороненными в курганах памятника Бертек-33. Обсуждаются вопросы корректного сопоставления и интерпретации данных о генетических характеристиках погребенных и особенностях материальной культуры исследуемого населения.

DNA sequencing reveals AMELX, ODAM and MMP20 variations in dental fluorosis

ObjectiveDental fluorosis (DF) is a dental development disorder caused by chronic fluoride overconsumption. There are differences in the susceptibility to and severity of DF in studied populations. The objective of the present study was to determine if single-nucleotide variations (SNVs) in the genes Amelogenin (AMELX), Odontogenic Ameloblast Associated (ODAM) and Matrix Metalloproteinase 20 (MMP20) are associated with DF by evaluating the relationship between variations in these genes and the degree of DF severity. Subjects and methodsSchoolchildren from two regions of Durango State and Mexico City, Mexico, were studied. The DF phenotype was determined using the Thylstrup and Fejerskov (TF) index. DNA was obtained from the buccal mucosa of each participant, and the presence of the variations rs946252 in AMELX, rs1514392 in ODAM and rs1784418 in MMP20 was determined by bidirectional DNA sequencing. ResultsA total of 180 DNA samples from 30 schoolchildren from 2 areas of Durango State were sequenced and analyzed. Differences in the severity of DF were found between the study areas (p = 0.006). SNVs in theMMP20 gene were present in 76.9 % of the participants in the high fluoride concentration and lower DF severity area. ConclusionAMELX and ODAM variations was not different between the two populations with respect to DF severity; however, the presence of rs1784418 differed between phenotypes with regard to susceptibility to DF. Therefore, MMP20 might be related to the various phenotypes of DF and may serve as a protective marker.

The Evolution of Unusually Small Amelogenin Genes in Cetaceans; Pseudogenization, X-Y Gene Conversion, and Feeding Strategy.

Among extant cetaceans, mysticetes are filter feeders that do not possess teeth and use their baleen for feeding, while most odontocetes are considered suction feeders, which capture prey by suction without biting or chewing with teeth. In the present study, we address the functionality of amelogenin (AMEL) genes in cetaceans. AMEL encodes a protein that is specifically involved in dental enamel formation and is located on the sex chromosomes in eutherians. The X-copy AMELX is functional in enamel-bearing eutherians, whereas the Y-copy AMELY appears to have undergone decay and was completely lost in some species. Consistent with these premises, we detected various deleterious mutations and/or non-canonical splice junctions in AMELX of mysticetes and four suction feeding odontocetes, Delphinapterus leucas, Monodon monoceros, Kogia breviceps, and Physeter macrocephalus, and in AMELY of mysticetes and odontocetes. Regardless of the functionality, both AMELX and AMELY are equally and unusually small in cetaceans, and even their functional AMELX genes presumably encode a degenerate core region, which is thought to be essential for enamel matrix assembly and enamel crystal growth. Furthermore, our results suggest that the most recent common ancestors of extant cetaceans had functional AMELX and AMELY, both of which are similar to AMELX of Platanista minor. Similar small AMELX and AMELY in archaic cetaceans can be explained by gene conversion between AMELX and AMELY. We speculate that common ancestors of modern cetaceans employed a degenerate AMELX, transferred from a decaying AMELY by gene conversion, at an early stage of their transition to suction feeders.

A rare genetic gender anomaly identified in a paternity case presenting AMEL-Y dropout

The examination of different sets of sex markers in view of gender determination may seem a routine and yet, no forensic geneticist is exempt from dealing with challenging cases on this issue, even if genetic anomalies related to gender occur rarely. We report a AMEL-Y dropout case revealed on the occasion of a paternity investigation, which proved to be a rare, 46,XX (SRY+) male syndrome, also known as de la Chapelle syndrome. The objective of our case report is to highlight the importance of forensic PCR kits to elucidate anomalies related to gender and overcome the problems related to a wrong genetic gender determination in paternity testing or human identification. The subject is a 36-year-old male requesting a paternity test because he was having doubts related to his 2 year-old child born during his marriage. The standard testing results performed on 16 STRs indicated an exclusion from paternity of our male subject who additionally exhibited an AMEL-Y dropout. We further investigated the male sample using a combination of PCR kits (Y-STRs, X-STRs, Amelogenin and SRY gene) and were able to identify the SRY gene, the loss of the major part of the Y-chromosome, except of a small region surrounding the SRY gene, and the presence of two X-chromosomes. Thus, we finally characterized the AMEL-Y anomaly as being a 46,XX male syndrome. Some basic facts related to the frequency, ethiopathology and diagnosis of this syndrome are reviewed and discussed. This study also represents the first reported Amelogenin dropout case detected in the Romanian population.

Cytogenetic Analyses in Ewes with Congenital Abnormalities of the Genital Apparatus.

Simple SummaryThe disorders of sex development (DSDs) are congenital conditions characterized by inconsistency among chromosomal, gonadal, and anatomical sex development. Reproduction and prolificacy are two main parameters in the sheep industry; thus, DSDs in sheep are very detrimental. Interestingly, no DSDs-associated gene mutations have been reported in sheep so far, probably due to the fact that affected animals are not detected and studied. With the aim to deepen the knowledge about DSDs in sheep and improve diagnostic tools, screening of a sheep farm aimed to detect and study DSDs in ovine species has been started, and the actual findings relative to the first two sheep flocks analysed are reported here. In our opinion, this study demonstrates that, despite the low number of studies on ovine DSDs, if compared with other species, this problem is actually present and needs more attention.The Disorders of Sex Development (DSDs) are congenital conditions characterized by inconsistency among chromosomal, gonadal, and anatomical sex development. The aim of this research is to report the clinical and cytogenetic findings of four DSD cases and 13 couples of heterosexual twins in sheep. To this purpose, C- and R-banding techniques were used, and the analyses of the SRY (Sex Determining Region Y) and AMEL (Amelogenin) genes were carried out. Moreover, morphopathological analyses were performed in one case. The four DSD sheep cases were registered as females at birth, and for none of them it was possible to establish whether the subjects were born from heterosexual multiple births. Three of the four cases were diagnosed as XX/XY blood lymphocyte chimaeras, while the fourth case was diagnosed as a 54, XY SRY-positive DSD sheep. None of the heterosexual twins showed XX/XY blood chimaerism. This finding suggests that the blood chimaeric cases detected could also be due to a zygote/embryo fusion. Moreover, no gene variants involved in sheep DSD are known, the identification of which would be very useful for the sheep industry.

Genetic diversity and phylogenetic relationships in feral pig populations from Argentina

In Argentina, domestic pigs (Sus scrofa Linnaeus 1758) were introduced during the first Buenos Aires foundation, in the year 1536. Their provenance was mainly from the Iberian Peninsula, the Canary Islands and Cape Verde. In 1541 those pigs were released and, consequently, the first feral populations were originated. Thereafter, the species propagated both naturally and through human action, reaching a distribution that covers most of the Argentinian territory. The objective of this study is to genetically characterize the oldest feral pig populations in Argentina, making use of the mitochondrial control region (CR) and the amelogenin gene (AmelY), in order to determine their phylogenetic origin and corroborate its consistency with the historic information. The obtained results indicate that most of the feral pigs in Corrientes and Buenos Aires populations are positioned in the European subclades, E1-A and E1-C for CR, and HY1 and HY2 for AmelY. Despite this fact, a low frequency of individuals of Asian origin was found in populations from Buenos Aires, whereas none of them disclosed African ancestry. Furthermore, given that a large proportion of feral pigs found in the species’ original sites in Argentina have European ancestry, we can partially corroborate the historical records.

Analysis of ancient mtDNA from the medieval archeological site of Amiternum (L'Aquila), central Italy

Study of ancient DNA makes it possible to analyze genetic relationships between individuals and populations of past and present. In this paper we have analyzed remains of human bones, dating back to the 8th-10th century AD, from the burials found in the Cathedral of Santa Maria in Civitate, archaeological site of Amiternum, L'Aquila, Italy. As a genetic marker, the hypervariable region 1 of mitochondrial DNA (HVR1) was selected. To obtain reliable sequences from the hypervariable region 1 of mtDNA (HVR1) were performed: multiple extractions, template quantification and cloning of PCR products. The sequences obtained were compared with Anderson's sequence for the identification of polymorphisms (SNP) and haplogroups. The data obtained were analyzed with various software and phylogenetic methods. For the comparison between populations, ancient and modern sequences found in databases and literature have been used. This work provides preliminary information on the correlation between the population of Amiternum, the migrant populations transited and/or established in the territory of Amiternum such as Byzantines, Longobards (Lombards), which dominated the Italian peninsula between 568 and 774 AD, and the current populations of Italy.The study of haplogroups, the analysis of genetic variability and phylogenesis studies on the sequences considered show a genetic closeness between the individuals of Amiternum, the current population of central-northern Italy and the Germanic tribe of Longobards, however, also highlights genetic traits of Byzantines in some samples of Amiternum. Using the analysis of amelogenin gene fragments, we successfully determined the sex of the bone remains on all samples.

Generation of Amelx-iCre Mice Supports Ameloblast-Specific Role for Stim1

The identification and targeting of the molecular pathways regulating amelogenesis is an ongoing challenge in dental research, and progress has been restricted by the limited number of genetic tools available to study gene function in ameloblasts. Here, we generated 4 transgenic Cre-driver mouse lines that express improved Cre (iCre)–recombinase from the locus of the mouse ameloblast-specific gene amelogenin X (Amelx-iCre) with a large (250-kb) bacterial artificial chromosome DNA vector. All 4 Amelx-iCre transgenic lines were bred with ROSA26 reporter mice to characterize the iCre developmental pattern with the LacZ gene encoding β-galactosidase enzyme activity assay and Cre protein immunohistochemistry. From the 4 generated transgenic lines, 2 were selected for further analysis because they expressed a high amount of Cre recombinase exclusively in ameloblasts and showed developmental stage- and cell-specific β-galactosidase activity mimicking the endogenous amelogenin expression. To test the functionality of the selected transgenic models, we bred the 2 Amelx-iCre mice lines with stromal interaction molecule 1 (Stim1) floxed mice to generate ameloblast-specific Stim1 conditional knockout mice (Stim1 cKO). STIM1 protein serves as one of the main calcium sensors in ameloblasts and plays a major role in enamel mineralization and ameloblast differentiation. Amelx-iCre mice displayed exclusive CRE-mediated recombination in incisor and molar ameloblasts. Stim1 cKO mice showed a severely defected enamel phenotype, including reduced structural integrity concomitant with increased attrition and smaller teeth. The phenotype and genotype of the Amelx-iCre/Stim1 cKO showed significant differences with the previously reported Ker14-Cre/Stim1 cKO, highlighting the need for cell- and stage-specific Cre lines for an accurate phenotype-genotype comparison. Furthermore, our model has the advantage of carrying the entire Amelx gene locus rather than being limited to an Amelx partial promoter construct, which greatly enhances the stability and the specificity of our Cre expression. As such, the Amelx-iCre transgenic lines that we developed may serve as a powerful tool for targeting ameloblast-specific gene expression in future investigations.

Full-sibling allelic frequency and sharing among Madurese: STR technique by 12 locus and the sex-typing amelogenin gene

BackgroundSibling relation over chosen STR loci and localized database is yet to be developed in Indonesia like into many other countries despite the demand of varied ethnic population, catastrophes and civil conflict that call forensic preparedness. The targeted application includes human genomic typing and profile, kinship/sibship relation, forensic applications, individual identification, and or claneology tracing. Consequently, this study devotes on sibship in assessing and establishing preferable STR loci and allelic sharing extent amongst Madurese full sibling.Materials and methodsOne hundred blood samples were used to examine sibling relationship among 50 full siblings (25 pairs) assigned from a total of 100 individuals of 25 Madura families. Sibship relation was done by STR technique profile by 12 loci (CSF1PO, F13B, FES, TH01, TPOX, vWA, D5S818, D7S820, D8S1179, D13S317, D16S539, and D18S51) and a sex-typing amelogenin gene locusResultsFrom 300 observations (25 pairs of full-sibling × 12 loci) of genotyped STR allele, of allele sharing, has found 4 STR loci: FES, CSF1PO, F13, and D8S1179 with high-shared allelic frequency in two alleles sharing among twelve STR loci by 52%, 52%, 44%, and 40% respectively. As for the four main loci of full sibling with one allele sharing, i.e, D7S820, D18S51, vWA, and THO1, have a sequence of percentages from large to small as follows: 76%, 76%, 72%, and 60%, respectively. Furthermore, STR locus VWA expressed null shared allele in 24 allele-sharing category. Likewise, F13 and FES both typed null allele in zero allele-sharing category among these full siblingConclusionThrough the use of 12 autosomal STR loci, the study has signified the strength of 2 shared allele evidence by 51.2% in establishing sibship and recommend D7S820, VWA, TH01, and D18S51 as STR loci of choice when typing sibling among Madurese.

Development of a new 25plex STRs typing system for forensic application.

We have developed a novel STR 25-plex florescence multiplex-STR kit (DNATyper25) to genotype 23 autosomal and two sex-linked loci for forensic applications and paternity analysis. Of the 23 autosomal loci, 20 are non-CODIS. The sex-linked markers include a Y-STR locus (DYS391) and the Amelogenin gene. We present developmental validation studies to show that the DNATyper25 kit is reproducible, accurate, sensitive, and robust. Sensitivity testing showed that full profiles were achieved with as low as 125pg of human DNA. Specificity testing demonstrated a lack of cross reactivity with a variety of commonly encountered non-human DNA contaminants. Stability testing showed that full profiles were obtained with humic acid concentration ≤60ng/μL and hematin concentration <400μM. For forensic evaluation, the 23 autosomal STRs followed the Hardy-Weinberg equilibrium. In an analysis of 509 Chinese (CN) Hans, we detected a combined total of 181 alleles at the 23 autosomal STR loci. Since these autosomal STRs are independent from one another, PM was 8.4528×10-22 , TDP was 0.999 999 999 999 999 999 999, CEP was 0.999 999 8395. The forensic efficiency parameters demonstrated that these autosomal STRs are highly polymorphic and informative in the Han population of China. We performed population comparisons and showed that the Northern CN Han has a close genetic relationship with the Luzhou Han, Tujia, and Bai populations. We propose that the DNATyper25 kit will be useful for cases where paternity analysis is difficult and for situations where DNA samples are limited in quantity and low in quality.

Analysis of the Yp11.2 Deletion Region of Phenotypically Normal Males with an AMELY-Null Allele in the Chinese Han Population

Aim: The amelogenin gene is a widely used gender marker for forensic DNA profiling. Males who have the amelogenin Y (AMELY) allele deletion can be mistakenly identified as females if genotyping is performed only on the amelogenin gene. The aim of this study was to investigate the frequency of the AMELY allele deletion in the Chinese Han population and analyze the possible genetic variation on the Y chromosome. Materials and Methods: The amelogenin gene of 12,735 unrelated males from the Chinese Han population were genotyped using common forensic short tandem repeat (STR) kits. The AMELY allele deletion was verified by redesigned primers and sequencing. Eighteen Y-specific sequence tagged sites (STSs) on the Yp11.2 region were selected to delineate the deletion breakpoints on the Y chromosome. Results: Three males were confirmed to have no AMELY allele. The frequency rate of the AMELY-null allele was 0.236% (3/12,735) in the Chinese Han population of Central China; 2.73 Mb of sequence on the Y chromosome were absent in all the AMELY-negative samples. The deleted region was mapped using SRY, AMELY, 5 Y-STRs, and 18 STSs, which belong to the class I deleted pattern. The three unrelated males shared the same Y-STR haplotype with four males from other Chinese populations, all of whom have the AMELY-null allele. The haplogroup of these males was identified as the O3 haplogroup. Conclusion: The AMELY allele deletion in the Chinese population was accompanied by the deletion of the Y-STR loci on the Yp11.2 region. Therefore, another Y-specific marker should be tested simultaneously when unknown samples are examined as part of a criminal investigation.

Genetic diversity and forensic characteristics of 15 autosomal short tandem repeats in Tibet highland and Guangdong lowland Han Chinese populations

Background: Much attention has been paid to the genetic variants of microsatellites in East Asian populations; however, genetic diversity of high-altitude adaptation in Tibet Han remains largely uncharacterised.Aim: To profile DNA samples from 338 high-altitude adaptation Han Chinese individuals and 933 low-altitude living Han Chinese individuals for 15 autosomal STRs which are used for human identification purposes and to estimate the forensic parameters as well as explore the genetic relationships among 38 Chinese populations.Subjects and methods: Fifteen autosomal STR loci and amelogenin genes were amplified in 1271 individuals using the AmpFℓSTR® Sinofiler™ PCR Amplification Kit. Allele frequencies and forensic parameters were calculated. Subsequently, population comparisons among 38 groups were analysed via principal components analysis, Reynolds genetic distance, neighbour-joining tree and multidimensional scaling plots.Results: In this study, no departures from Hardy-Weinberg equilibrium are identified in either Tibet or Guangdong Han populations after Bonferroni correction. The cumulative match probabilities are 3.1108 × 10−19 in Guangdong Han and 6.2102 × 10−19 in Tibet Han, and the combined probabilities of exclusion for trios are 0.99999948 and 0.99999936, respectively. Comprehensive population comparisons based on allele frequency distribution indicate that the Tibet Han population has a genetically close relationship with the surrounding population (Tibet Tibetan) and the Guangdong Han population has genetic affinity with southern Chinese populations.Conclusion: In general, genetic polymorphisms and forensic efficiency indicated that the 15 STRs studied are informative and polymorphic in both lowland and highland Han populations.

Establishing gene Amelogenin as sex-specific marker in yak by genomic approach

Yak, an economically important bovine species considered as lifeline of the Himalaya. Indeed, this gigantic bovine is neglected because of the scientific intervention for its conservation as well as research documentation for a long time. Amelogenin is an essential protein for tooth enamel which eutherian mammals contain two copies in both X and Y chromosome each. In bovine, the deletion of a fragment of the nucleotide sequence in Y chromosome copy of exon 6 made Amelogenin an excellent sex-specific marker. Thus, an attempt was made to use the gene as an advanced molecular marker of sexing of the yak to improve breeding strategies and reproduction. The present study confirmed that the polymerase chain reaction amplification of the Amelogenin gene with a unique primer is useful in sex identification of the yak. The test is further refined with qPCR validation by quantifying the DNA copy number of the Amelogenin gene in male and female. We observed a high level of sequence polymorphisms of AMELX and AMELY in yak considered as novel identification. These tests can be further extended into several other specialized fields including forensics, meat production and processing, and quality control.

Identification of Amelogenin Gene on Burnt Teeth Samples through Nested Polymerase Chain Reaction Amplification for Sex Identification

Sex determination is one of the basic components in victim identification. This study aims to ascertain the sex of an individual from burnt teeth samples exposed at different temperature and time through nested polymerase chain reaction (PCR) on the amelogenin (AMEL) sex marker, to calculate the specificity and sensitivity, and to compare with previous relevant studies. A total of 17 teeth samples was subjected to burning at different temperatures ranging from 100°C to 500°C, at 2 to 10 minutes. The whole tooth was used for deoxyribonucleic acid (DNA) extraction by phenol-chloroform method. All samples were quantified for DNA concentration and then analyzed with nested PCR using two pairs of AMEL primer and results of sex typing were recorded. Out of 17 samples, genomic DNA extracted from 6 samples have concentrations ranging from 27.3 – 130.6 ng/µL. Nested PCR could amplify 16 samples for AMEL gene. Sex typing using AMEL gene showed 76.47% accuracy. Sensitivity of AMEL primer was increased from 6.67% to 63.64% using nested PCR technique; specificity of both external and internal primer was reported at 100%. Nested PCR of AMEL gene proved to be a suitable method for unequivocal determination of sex from degraded DNA samples. DOI : http://dx.doi.org/10.17576/JSKM-2019-1701-12

Molecular-genetic analysis of a rare forensic case of partial deletion of the Y-chromosome

The amelogenin gene encodes dental enamel protein and is present in humans in two forms - AMELX and AMELY, located on the X- and Y-chromosomes, respectively. This rare case depicts a partial deletion of the AMELY gene. In the Into-Stil LLC laboratory, we performed the genetic testing of the DNA samples extracted from buccal epithelial cells of the alleged father and the disputed child (a boy). Genotyping was carried out using COrDIS Plus ('Gordis', Russian Federation) and AmpFLSTR Identifiler Direct PCR Amplification ('Applied Biosystems', USA) Kits. Our findings have demonstrated that both the alleged father and the disputed child lacked the fragments corresponding with the AMELY gene. Using both STR-systems, we detected, in the disputed child's genome, the allele formally identical to the allele in the genome of the alleged father. Further analysis using the COrDYS ('Gordis', Russian Federation) kit allowed us to detect the amplified fragments corresponding with all the STR loci of Y chromosome, except DYS576 and DYS449, which confirmed that both studied individuals belonged to male biological sex.