Browsing by Author "Stojković, Oliver (35618950700)"

Myotonic dystrophy type 1 (DM1) is associated with an expansion of CTG repeats in the 3′UTR of the DMPK gene. It is accepted, as in other trinucleotide diseases, that the number of the repeats is correlated with age at onset and severity of the disease. However, assessment of genotype-phenotype correlation in DM1 is complicated with the expansion-biased somatic instability of mutant alleles over time and difficulties in precise assessment of the number of repeats by standard Southern blot hybridization. In order to clarify this issue we defined DM1 expansion size in lymphocytes by three parameters: size of progenitor, average, and largest allele, using a more precise small-pool/long-range PCR technique. We found a negative linear correlation of age at onset and average expansion size in juvenile-adult DM1 patients (35 out of 46) whose progenitor allele is less than 245 repeats long. Our result favors the hypothesis of the existence of a threshold in the progenitor allele size beyond which number of CTG repeats does not influence age at onset. Potential clinical significance is that the average allele size could be a useful indicator for the age at onset in juvenile-adult DM1 patients with relatively short progenitor allele. To test whether somatic instability of mutant alleles influences the progression of DM1, patients were divided in three phenotypic classes according to the severity of neuromuscular symptoms. We showed that the largest expansion in each DM1 phenotypic class reflects somatic instability of mutant allele over time independently of progenitor allele size and patient's age at sampling. The mean of the largest expansion was significantly different between phenotypic classes, implying the possible association between expansion-biased somatic instability of mutant alleles over time and progression of neuromuscular symptoms. © 2002 Wiley-Liss, Inc.

Myotonic dystrophy type 1 (DM1) is associated with an expansion of CTG repeats in the 3′UTR of the DMPK gene. It is accepted, as in other trinucleotide diseases, that the number of the repeats is correlated with age at onset and severity of the disease. However, assessment of genotype-phenotype correlation in DM1 is complicated with the expansion-biased somatic instability of mutant alleles over time and difficulties in precise assessment of the number of repeats by standard Southern blot hybridization. In order to clarify this issue we defined DM1 expansion size in lymphocytes by three parameters: size of progenitor, average, and largest allele, using a more precise small-pool/long-range PCR technique. We found a negative linear correlation of age at onset and average expansion size in juvenile-adult DM1 patients (35 out of 46) whose progenitor allele is less than 245 repeats long. Our result favors the hypothesis of the existence of a threshold in the progenitor allele size beyond which number of CTG repeats does not influence age at onset. Potential clinical significance is that the average allele size could be a useful indicator for the age at onset in juvenile-adult DM1 patients with relatively short progenitor allele. To test whether somatic instability of mutant alleles influences the progression of DM1, patients were divided in three phenotypic classes according to the severity of neuromuscular symptoms. We showed that the largest expansion in each DM1 phenotypic class reflects somatic instability of mutant allele over time independently of progenitor allele size and patient's age at sampling. The mean of the largest expansion was significantly different between phenotypic classes, implying the possible association between expansion-biased somatic instability of mutant alleles over time and progression of neuromuscular symptoms. © 2002 Wiley-Liss, Inc.

In Europe, the first case of coronavirus disease (COVID-19) and the first COVID-19-related death were reported in France on January 24th and February 15th, 2020, respectively. Officially, the first case of COVID-19 infection in the Republic of Serbia was registered on March 6th. Herein, we presented the first case of retrospective detection of the Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) in the post-mortem-obtained vitreous humor (VH), which took place on February 5th, 2020. This is the first death in Europe proven to be caused by COVID-19 by means of post-mortem histopathological and molecular analyses. Based on this finding, it appears that SARS-CoV-2 has been spreading faster and started spreading much earlier than it had been considered and that COVID-19 was probably the cause of the much-reported pneumonia of unknown origin in January and February 2020. © Copyright © 2021 Bogdanović, Skadrić, Atanasijević, Stojković, Popović, Savić, Mihailović, Radnić, Aćimović, Damjanjuk, Despotović and Barać.

Plethora of drugs and toxic substances is metabolized by cytochrome P450 enzymes (CYP450). These enzymes are coded by highly variable genes abundant with single nucleotide variants (SNVs) and small insertions/deletions (indels) that affect the functionality of the enzymes, increasing or decreasing their activity. CYP genes genotyping, followed by haplotype inference, provides substrate specific metabolic phenotype prediction. This is crucial in pharmacogenetics and applicable in molecular autopsy. However, high number of alleles in CYP450 superfamily and interethnic variability in frequency distribution require precise gene panel customization. To estimate informativeness of SNVs and alleles in CYP gene families 1, 2, and 3, associated with metabolic alterations, 500 unrelated individuals from 5 regions of Serbia were genotyped using TaqMan assays to determine frequencies of CYP2C9 *2 and *3, CYP2C19 *2 and *17 alleles, four variants in CYP2D6 (rs3892097, rs1065852, rs28371725, rs28371706) gene, and CYP3A4*1B allele. In addition, CYP1A1 rs4646903 and rs1048943 (m1 and m2) variants were genotyped by RFLP. Our results showed that frequencies of tested variants in Serbian population corresponded to general European population and somewhat differed from neighboring populations. SNV rs1065852, the main contributor to non-functional CYP2D6 *4, significantly departed from Hardy-Weinberg equilibrium. With the exception of rs28371706 in CYP2D6 and rs2740574 in CYP3A4, which were very rare in our sample, all other tested variants in CYP2 family are informative and appropriate for pharmacogenetic testing, molecular autopsy, and medico-legal genetic analyses. © 2019, Springer-Verlag GmbH Germany, part of Springer Nature.

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Human-specific quantitative PCR (qPCR) has been developed for forensic use in the last 10 years and is the preferred DNA quantification technique since it is very accurate, sensitive, objective, time-effective and automatable. The amount of information that can be gleaned from a single quantification reaction using commercially available quantification kits has increased from the quantity of nuclear DNA to the amount of male DNA, presence of inhibitors and, most recently, to the degree of DNA degradation. In skeletal remains samples from disaster victims, missing persons and war conflict victims, the DNA is usually degraded. Therefore the new commercial qPCR kits able to assess the degree of degradation are potentially able to predict the success of downstream short tandem repeat (STR) typing. The goal of this study was to verify the quantification step using the PowerQuant kit with regard to its suitability as a screening method for autosomal STR typing success on ancient and Second World War (WWII) skeletal remains. We analysed 60 skeletons excavated from five archaeological sites and four WWII mass graves from Slovenia. The bones were cleaned, surface contamination was removed and the bones ground to a powder. Genomic DNA was obtained from 0.5 g of bone powder after total demineralization. The DNA was purified using a Biorobot EZ1 device. Following PowerQuant quantification, DNA samples were subjected to autosomal STR amplification using the NGM kit. Up to 2.51 ng DNA/g of powder were extracted. No inhibition was detected in any of bones analysed. 82% of the WWII bones gave full profiles while 73% of the ancient bones gave profiles not suitable for interpretation. Four bone extracts yielded no detectable amplification or zero quantification results and no profiles were obtained from any of them. Full or useful partial profiles were produced only from bone extracts where short autosomal (Auto) and long degradation (Deg) PowerQuant targets were detected. It is concluded that STR typing of old bones after quantification with the PowerQuant should be performed only when both Auto and Deg targets are detected simultaneously with no respect to [Auto]/[Deg] ratio. Prediction of STR typing success could be made according to successful amplification of Deg fragment. The PowerQuant kit is capable of identifying bone DNA samples that will not yield useful STR profiles using the NGM kit, and it can be used as a predictor of autosomal STR typing success of bone extracts obtained from ancient and WWII skeletal remains. © 2016 Elsevier Ireland Ltd

Human-specific quantitative PCR (qPCR) has been developed for forensic use in the last 10 years and is the preferred DNA quantification technique since it is very accurate, sensitive, objective, time-effective and automatable. The amount of information that can be gleaned from a single quantification reaction using commercially available quantification kits has increased from the quantity of nuclear DNA to the amount of male DNA, presence of inhibitors and, most recently, to the degree of DNA degradation. In skeletal remains samples from disaster victims, missing persons and war conflict victims, the DNA is usually degraded. Therefore the new commercial qPCR kits able to assess the degree of degradation are potentially able to predict the success of downstream short tandem repeat (STR) typing. The goal of this study was to verify the quantification step using the PowerQuant kit with regard to its suitability as a screening method for autosomal STR typing success on ancient and Second World War (WWII) skeletal remains. We analysed 60 skeletons excavated from five archaeological sites and four WWII mass graves from Slovenia. The bones were cleaned, surface contamination was removed and the bones ground to a powder. Genomic DNA was obtained from 0.5 g of bone powder after total demineralization. The DNA was purified using a Biorobot EZ1 device. Following PowerQuant quantification, DNA samples were subjected to autosomal STR amplification using the NGM kit. Up to 2.51 ng DNA/g of powder were extracted. No inhibition was detected in any of bones analysed. 82% of the WWII bones gave full profiles while 73% of the ancient bones gave profiles not suitable for interpretation. Four bone extracts yielded no detectable amplification or zero quantification results and no profiles were obtained from any of them. Full or useful partial profiles were produced only from bone extracts where short autosomal (Auto) and long degradation (Deg) PowerQuant targets were detected. It is concluded that STR typing of old bones after quantification with the PowerQuant should be performed only when both Auto and Deg targets are detected simultaneously with no respect to [Auto]/[Deg] ratio. Prediction of STR typing success could be made according to successful amplification of Deg fragment. The PowerQuant kit is capable of identifying bone DNA samples that will not yield useful STR profiles using the NGM kit, and it can be used as a predictor of autosomal STR typing success of bone extracts obtained from ancient and WWII skeletal remains. © 2016 Elsevier Ireland Ltd