Browsing by Subject "genotyping"

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  • Benedek, P.; Jiao, H.; Duvefelt, K.; Skoog, T.; Linde, M.; Kiviluoma, P.; Kere, J.; Eriksson, M.; Angelin, B. (2021)
    Aim To investigate whether genotyping could be used as a cost-effective screening step, preceding next-generation sequencing (NGS), in molecular diagnosis of familial hypercholesterolaemia (FH) in Swedish patients. Methods and results Three hundred patients of Swedish origin with clinical suspicion of heterozygous FH were analysed using a specific array genotyping panel embedding 112 FH-causing mutations in the LDLR, APOB and PCSK9 genes. The mutations had been selected from previous reports on FH patients in Scandinavia and Finland. Mutation-negative cases were further analysed by NGS. In 181 patients with probable or definite FH using the Dutch lipid clinics network (DLCN) criteria (score >= 6), a causative mutation was identified in 116 (64%). Of these, 94 (81%) were detected by genotyping. Ten mutations accounted for more than 50% of the positive cases, with APOB c.10580G>A being the most common. Mutations in LDLR predominated, with (c.2311+1_2312-1)(2514)del (FH Helsinki) and c.259T>G having the highest frequency. Two novel LDLR mutations were identified. In patients with DLCN score < 6, mutation detection rate was significantly higher at younger age. Conclusion A limited number of mutations explain a major fraction of FH cases in Sweden. Combination of selective genotyping and NGS facilitates the clinical challenge of cost-effective genetic screening in suspected FH. The frequency of APOB c.10580G>A was higher than previously reported in Sweden. The lack of demonstrable mutations in the LDLR, APOB and PCSK9 genes in similar to 1/3 of patients with probable FH strongly suggests that additional genetic mechanisms are to be found in phenotypic FH.
  • Pietarinen, Paavo (Helsingfors universitet, 2012)
    Most xenobiotics are biotransformed by phase I enzymes to a more hydrophilic form in order to get excreted out from the body. In most cases xenobiotics are in lipophilic form when entering body. The most important group in phase I enzymes is cytochrome P450 (CYP) superfamily. Of CYP enzymes probably the most studied is CYP2D6, which is responsible for metabolism of 20-25% of drugs currently on market. Many CYP2D6 substrates belong to therapeutically important drug groups, such as antiarrhytmics, antidepressants, beta-blockers, or neuroleptics. CYP2D6 gene, which encodes the enzyme, exhibits large interindividual variability, which has an effect on the metabolic activity of the enzyme. The frequencies of these genetic variances differ globally on wide scale between and inside populations. Through genotyping it is possible to predict the CYP2D6 metabolic rate, which can be divided into four classes: ultra-rapid metabolizers (UM), extensive metabolizers (EM), intermediate metabolizers (IM), and poor metabolizers (PM). The purpose of our study was to examine the frequencies of CYP2D6 genotypes in Finnish population in detail and compare the results to previous studies. Our study population consisted of 857 healthy volunteers whose DNA was extracted. From DNA sample we genotyped 10 different CYP2D6 genetic variants and the copy number of the gene using Applied Biosystems TaqMan genotyping and copy number assays. This study was the largest CYP2D6 genotype frequency study in Finnish population so far. The results supported the findings of a similar study in a Finnish population of smaller scale. Large majority of study subjects were EMs (87.3%) and the second largest group was Ums (7.2%). IMs and PMs were in clear minority (3.0% and 2.5%, respectively). The expected frequencies for UMs (1-2%) are much lower and for PMs higher (~8%) in other North European populations than in Finns. Accordingly, CYP2D6 genetic profile of Finnish population differs from its neighbours, which may be important for the dose requirements, efficacy, and safety for drugs metabolized by CYP2D6.
  • Duplouy, Anne; Nair, Abhilash; Nyman, Toshka; van Nouhuys, Saskya (2021)
    Population bottlenecks associated with founder events strongly impact the establishment and genetic makeup of populations. In addition to their genotype, founding individuals also bring along symbionts that can manipulate the phenotype of their host, affecting the host population establishment, dynamics and evolution. Thus, to understand introduction, invasion, and spread, we should identify the roles played by accompanying symbionts. In 1991, the parasitoid wasp, Hyposoter horticola, and its associated hyperparasitoid were accidentally introduced from the main Åland islands, Finland, to an isolated island in the archipelago, along with their host, the Glanville fritillary butterfly. Though the receiving island was unoccupied, the butterfly was present on some of the small islands in the vicinity. The three species have persisted as small populations ever since. A strain of the endosymbiotic bacterium Wolbachia has an intermediate prevalence in the H. horticola across the main Åland population. The infection increases susceptibility of the parasitoid to hyperparasitism. We investigated the establishment and spread of the parasitoid, along with patterns of prevalence of its symbiont using 323 specimens collected between 1992 and 2013, from five localities across Åland, including the source and introduced populations. Using 14 microsatellites and one mitochondrial marker, we suggest that the relatively diverse founding population and occasional migration between islands might have facilitated the persistence of all isolated populations, despite multiple local population crashes. We also show local near-fixation of Wolbachia, where the hyperparasitoid is absent, and selection against infected wasp genotypes is relaxed.
  • Kalendar, Ruslan; Baidyussen, Akmaral; Serikbay, Dauren; Zotova, Lyudmila; Khassanova, Gulmira; Kuzbakova, Marzhan; Jatayev, Satyvaldy; Hu, Yin-Gang; Schramm, Carly; Anderson, Peter A.; Jenkins, Colin L. D.; Soole, Kathleen L.; Shavrukov, Yuri (2022)
  • Youssef, Omar; Almangush, Alhadi; Zidi, Yossra H. S.; Loukola, Anu; Carpen, Olli (2020)
    Background:Archived formalin-fixed paraffin-embedded (FFPE) specimens from nonmalignant tissues derived from cancer patients are a vast and potentially valuable resource for high-quality genotyping analyses and could have a role in establishing inherited cancer risk. Methods:We systematically searched PubMed, Ovid MEDLINE, and Scopus databases for all articles that compared genotyping performance of DNA from nonmalignant FFPE tissue with blood DNA derived from cancer patients irrespective of tumor type. Two independent researchers screened the retrieved studies, removed duplicates, excluded irrelevant studies, and extracted genotyping data from the eligible studies. These studies included, but were not limited to, genotyping technique, reported call rate, and concordance. Results:Thirteen studies were reviewed, in which DNA from nonmalignant FFPE tissues derived from cancer patients was successfully purified and genotyped. All these studies used different approaches for genotyping of DNA from nonmalignant FFPE tissues to amplify single nucleotide polymorphisms (SNPs) and to estimate of loss of heterozygosity. The concordance between genotypes from nonmalignant FFPE tissues and blood derived from cancer patients was observed to be high, whereas the call rate of the tested SNPs was not reported in all included studies. Conclusion:This review illustrates that DNA from nonmalignant FFPE tissues derived from cancer patients can serve as an alternative and reliable source for assessment of germline DNA for various purposes, including assessment of cancer predisposition.
  • Kastally, Chedly; Niskanen, Alina K.; Perry, Annika; Kujala, Sonja T.; Avia, Komlan; Cervantes, Sandra; Haapanen, Matti; Kesälahti, Robert; Kumpula, Timo A.; Mattila, Tiina M.; Ojeda, Dario I.; Tyrmi, Jaakko S.; Wachowiak, Witold; Cavers, Stephen; Kärkkäinen, Katri; Savolainen, Outi; Pyhäjärvi, Tanja (2022)
    Pinus sylvestris (Scots pine) is the most widespread coniferous tree in the boreal forests of Eurasia, with major economic and ecological importance. However, its large and repetitive genome presents a challenge for conducting genome-wide analyses such as association studies, genetic mapping and genomic selection. We present a new 50K single-nucleotide polymorphism (SNP) genotyping array for Scots pine research, breeding and other applications. To select the SNP set, we first genotyped 480 Scots pine samples on a 407 540 SNP screening array and identified 47 712 high-quality SNPs for the final array (called 'PiSy50k'). Here, we provide details of the design and testing, as well as allele frequency estimates from the discovery panel, functional annotation, tissue-specific expression patterns and expression level information for the SNPs or corresponding genes, when available. We validated the performance of the PiSy50k array using samples from Finland and Scotland. Overall, 39 678 (83.2%) SNPs showed low error rates (mean = 0.9%). Relatedness estimates based on array genotypes were consistent with the expected pedigrees, and the level of Mendelian error was negligible. In addition, array genotypes successfully discriminate between Scots pine populations of Finnish and Scottish origins. The PiSy50k SNP array will be a valuable tool for a wide variety of future genetic studies and forestry applications.
  • Ripatti, Pietari (Helsingfors universitet, 2016)
    Familial combined hyperlipidemia (FCH) is a complex and common familial dyslipidemia characterized by elevated total cholesterol and/or triglyceride levels with over five-fold risk of coronary heart disease. The genetic architecture and contribution of rare Mendelian and common variants to FCH susceptibility is unknown. In 53 Finnish FCH families, we genotyped and imputed nine million variants in 715 family members with DNA available. We studied the enrichment of variants previously implicated with monogenic dyslipidemias and/or lipid levels in the general population by comparing allele frequencies between the FCH families and population samples. We also constructed weighted polygenic scores using 212 lipid-associated SNPs and estimated the relative contributions of Mendelian variants and polygenic scores to the risk of FCH in the families. We identified, across the whole allele frequency spectrum, an enrichment of variants known to elevate, and a deficiency of variants known to lower LDL-C and/or TG levels among both probands and FCH affecteds. The score based on TG associated SNPs was particularly high among affected individuals compared to non-affected family members. Out of 234 FCH affecteds across the families, seven (3 %) carried Mendelian variants and 83 (35 %) showed high accumulation of either known LDL-C or TG elevating variants by having either polygenic score over the 90th percentile in the population. There was large between-family variation in how much the polygenic scores contributed to the FCH phenotype. FCH is highly polygenic, supporting the hypothesis that variants across the whole allele frequency spectrum contribute to this complex familial trait. This reinforces the clinical tenet that FCH is a cluster of overlapping genetic defects instead of an etiologically homogenous disease entity.
  • Voutilainen, Miko (Helsingin yliopisto, 2022)
    Single nucleotide polymorphism (SNP) is DNA variation of a single nucleotide. SNPs are the most common mutations and millions of single nucleotide differences distinguish humans genetically from each other. Different gene variants of a single nucleotide affect nutritional and pharmacological metabolism and gene test results can therefore guide the diet and medication. SNPs are tested either by sequencing, gene arrays or quantitative polymerase chain reaction (qPCR). In this thesis the SNPs were tested by using 132 different TaqMan probes and the SmartChip qPCR platform. 20 participants donated two buccal swab samples and one dried blood spot (DBS) card. Buccal swabs were extracted using two different methods. DNA was also sent to a reference laboratory to be analysed by gene arrays and two participants also sent their DNA samples to two direct-to-consumer (DTC) commercial gene test companies. Low concentration buccal swab samples (<3 ng/µl) produced mismatches whereas DBS samples had high genotyping accuracy even at low DNA concentration. For buccal swabs there was a correlation between the DNA concentration and qPCR call rate (p<0.0001) but not for DBS samples (p>0.05). The allele separation of one TaqMan assay was not sufficient between minor homozygote and heterozygote clusters. For other 131 TaqMan assays excluding the low DNA concentration swab samples, the reproducibility rate was 99.97 %. The reproducibility rate with the DTC companies was 98.36 %. The lower reproducibility rate emphasizes the importance of the manual review of the genotyping raw data, which is not possible with huge sequencing or microarray SNP datasets. SNPs are found on various genetic structures which leads to inconsistent genotyping data that is difficult to analyse with a single algorithm. The SmartChip system combined with the TaqMan assays is a highly reproducible SNP genotyping method when the DNA concentration of the samples is high and the results are manually reviewed.