212 research outputs found
Computational Estimation of Fetal DNA Fraction in Low Coverage Whole Genome Sequencing Data
No full textKäesoleva töö eesmärk oli leida ja kalibreerida arvutuslik metoodika loote rakuvaba DNA fraktsiooni määramiseks raseda naise vereproovis. Tegemist on rakendusliku teadusuuringuga, mis on eeltingimuseks NIPT testi usaldusväärseks rakendamiseks tervishoiusüsteemis. NIPT on kõige täpsem ja kaasaegsem mitteinvasiivne loote sünnieelne kromosoomihaiguste sõeluuring, mis põhineb madala katvusega täisgenoomi sekveneerimisandmete analüüsil. Metoodika võimaldab määrata loote rakuvaba DNA hulka raseda naise vereproovis nii poiss- kui ka tüdruklootele, mis on vajalik, et iga raseduse rakuvaba DNA analüüsi tulemus oleks usaldusväärne ja arstile ning patsiendile edastatud tulemus tõene. Käesolevas töös kasutati madala katvusega üle-genoomsetest Illumina platvormiga läbiviidud sekveneerimise katsetest saadud 416 Eesti päritolu naise NIPT proove, et välja töötada Y-kromosoomi põhine loote rakuvaba DNA hulga määramine poiss-loodetele. Välja töötatud Y-kromosoomi põhist meetodit kasutati SeqFF arvutusliku metoodika valideerimiseks Eesti NIPT proovidel. SeqFF rakendamine Eesti NIPT proovidel võimaldab määrata loote rakuvaba DNA hulka nii poiss- kui ka tüdrukloodetel. Väljatöötatud algoritm on integreeritud Eestis pakutavasse NIPT täppismeditsiini teenusesse NIPTIFY.The aim of this thesis was to find and 'calibrate' computational methodology for estimating the proportion of cell-free fetal DNA (cffDNA) in pregnant women’s blood sample. This work was done as part of an applied research project aimed to develop a whole genome sequencing (WGS) based non-invasive prenatal testing (NIPT) medical screening test. NIPT is the most up-to-date, accurate and easily applied (non-invasive) prenatal screening method to detect fetal aneuploidies (for example trisomy 21, that causes Down syndrome) with high confidence and already during the first trimester of pregnancy. Commonly, NIPT is based on the low coverage WGS data, generated by the means of Illumina or some another platform technology. Computational tools used for aneuploidy detection can also estimate the proportion of cffDNA in maternal blood for both male and female fetus pregnancies. Fetal fraction calculation is a prerequisite to assure the technical credibility of NIPT screening test. In the current study, low coverage cell-free whole genome sequencing data from 416 pregnant women were used to develop a chromosome Y based estimator for the proportion of cffDNA in male-fetus pregnancy cases. Next, the chromosome Y based estimator was used to validate the credibility of SeqFF computational method with Estonian NIPT samples. This developed approach using SeqFF method on Estonian NIPT samples enables to estimate the proportion of cffDNA in both male and female fetus pregnancies. The SeqFF method is now integrated into the NIPT computation workflow service, validated and in daily practical use as part of the NIPTIFY® screening test
Arvutuslikud metoodikad loote trisoomiate ja mikrodeletsioonide riski tuvastamiseks mitteinvasiivses sõeluuringus
No full textVäitekirja elektrooniline versioon ei sisalda publikatsiooneMitteinvasiivne sünnieelne geneetiline testimine (NIPT) on sõeluuringu meetod loote kromosoomhaiguste riski hindamiseks raseda vereproovist. NIPT põhineb platsenta päritolu rakuvaba DNA sekveneerimisel ja andmeanalüüsil. Lisaks loote kromosoomi koopiaarvu muutustele võimaldab NIPT tuvastada ka patogeensete mikrodeletsioonide riski. Mikrodeletsiooni sündroom on lühikese kromosoomiosa kaost põhjustatud kromosoomihaigus, mille kliiniline raskusaste sõltub deleteerunud regioonist. Näiteks 22q11 piirkonnas esinev mikrodeletsioon põhjustab DiGeorge’i sündroomi, mis on seotud südamerikete, huule-suulaelõhe ja vaimupuudega. Loote kromosoomhaiguse riski määramine on bioinformaatiline väljakutse, sest miljonite DNA järjestuste korrektne analüüs eeldab nutikaid arvutuslikke lahendusi.
Käesoleva doktoritöö eesmärk on tõsta NIPT sünnieelse sõeluuringu täpsust, terviseandmete väärindamist ja meditsiiniteenuse kulutõhusust. Doktoritöö käigus loodi uus arvutuslik raamistik NIPT-i tarbeks ning valideeriti ja analüüsiti kliinilistel andmetel varasemalt publitseeritud NIPT algoritme. Lisaks töötati välja ja valideeriti kliiniliselt uudne tarkvarapakett BinDel, mis võimaldab hinnata loote mikrodeletsioonide riski.
Töö tulemusel kvantifitseeriti arvutuslike tööriistade täpsus erineva sekveneerimiskatvuse ja loote DNA osakaalu tingimustes. Tulemused näitasid, et NIPT täpsust mõjutab nii sekveneerimissügavus kui ka loote DNA osakaalu ning algoritmi valik. Uus BinDel tarkvara parandas mikrodeletsioonide tuvastamise võimekust, pakkudes võimalusi täpsemaks ja laialdasemaks sünnieelseks sõeluuringuks. Lisaks töötati välja masinõppepõhine arvutuslik raamistik loote kromosoomi koopiaarvu muutuste tuvastamiseks.Non-invasive prenatal genetic testing (NIPT) is a screening method for assessing the risk of fetal chromosomal disorders from a maternal blood sample. NIPT is based on sequencing and data analysis of cell-free DNA originating from the placenta. In addition to detecting changes in the fetal chromosome copy number, NIPT can also identify the risk of pathogenic microdeletions. A microdeletion syndrome results from the loss of a small segment of a chromosome, and its clinical severity depends on the deleted region. For example, a microdeletion in the 22q11 region causes DiGeorge syndrome, which is associated with heart defects, cleft palate, and intellectual disabilities. Fetal chromosomal disorder risk assessment is a bioinformatics challenge, as the accurate analysis of millions of DNA sequences requires sophisticated computational solutions.
The aim of this doctoral dissertation is to improve the accuracy, data utilization, and cost-effectiveness of NIPT prenatal screening. A new computational framework for NIPT was developed, and previously published NIPT algorithms were validated and analyzed using clinical data. Additionally, a novel software tool, BinDel, was developed and clinically validated for estimating fetal microdeletion risk.
The results quantified the accuracy of computational tools under varying sequencing coverage and fetal DNA fraction levels. The findings showed that NIPT accuracy is influenced by both sequencing depth and the fetal DNA fraction, as well as the choice of algorithm. The new BinDel software improved the ability to detect microdeletions, providing potential for more accurate and widespread prenatal screening. Additionally, a machine learning-based computational framework was developed for detecting fetal chromosomal copy number changeshttps://www.ester.ee/record=b575094
Arvutuslikud meetodid DNA koopiaarvu määramiseks
No full textVäitekirja elektrooniline versioon ei sisalda publikatsioone.DNA koopiaarvu variantideks või muutusteks nimetatakse selliseid erinevusi inimeste geneetilises materjalis, mille puhul mingi DNA lõigu koopiaarv on erinev oodatavast koopiaarvust kaks (üks koopia mingit kindlat DNA järjestust emalt päritud kromosoomil ja üks koopia isalt päritud kromosoomil). DNA koopiate vähenemist nimetatakse deletsiooniks ning vastavaid DNA variante nimetatakse deletsioonideks. DNA koopiate juurdetulemist nimetatakse duplitseerumiseks ning selliseid kahest suurema koopiaarvuga variante vastavalt duplikatsioonideks.
Antud doktoritöös uuriti inimese DNA koopiaarvu variante, nende seotust erinevate haigustega ja nende tekkimise ja pärandumise eripärasid. Kasutades DNA mikrokiipe ehk geenikiipe uuriti esmalt kas ja millised DNA koopiaarvu muutused võivad olla seotud vaimse arengu mahajäämusega (VAM-ga). Uurides perekondasid, kus ühel või mitmel liikmel oli diagnoositud VAM, leiti mitmeid juba varem VAM-ga seostatud DNA koopiaarvu muutusi ning lisaks leiti ka mitmeid uusi DNA koopiaarvu variante, mille esinemine võib olla seotud VAM-e väljakujunemisega.
Sarnane uuring viidi läbi ka korduva spontaanse raseduse katkemise probleemiga paaride ja naiste puhul. Võrreldes nende patsientide gruppi kuuluvate naiste DNA koopiaarvu muutusi ning nende sagedusi terveid emasid sisaldavate kontroll-grupi indiviidide omadega, leiti statistiliselt ja bioloogiliselt oluline erinevus muutunud koopiaarvuga DNA lõigus, mis sisaldab PDZD2 ja GOLPH3 geene ja kus esinevate duplikatsioonide „omamine“ suurendas naistel märkimisväärselt spontaanse raseduse katkemise ohtu.
Doktoritöö viimases osas uuriti Tartu Ülikooli Eesti Geenivaramu ja rahvusvahelise HapMap projekti poolt kogutud tõsiste haigusteta inimestel esinevaid DNA koopiaarvu muutusi ja nende pärandumist perekondades. Selle uuringu üheks huvitavamaks tulemuseks oli deletsioonide alapärandumine vanematelt lastele ehk deletsioone kandvaid DNA regioone esines laste genoomides oluliselt vähem, kui normaalse Mendeliaalse (juhusliku) pärandumise korral oleks oodata võinud. Uurides duplikatsioonide regioone perekondades leiti aga, et kaks kolmandikku duplikatsioonides esinevatest DNA koopiatest ei olnud identsed (üksteise täpsed koopiad), vaid mõnevõrra erinevad, demonstreerides seniajani teadmata olnud alleelse varieeruvuse määra DNA duplikatsioonide regioonides.DNA copy number variation is a type of genetic variation in which case the number of copies of a particular region of a chromosome is altered from its normal state. In the non-repetitive portion of the human genome, the normal haploid copy number is one – one copy of each sequence per chromosome. Accordingly, the normal diploid copy number in humans is two – one copy inherited from both parents. A copy number variant (CNV) can result from either a loss of copies (most often called a deletion) or gain of copies (called a duplication or amplification).
In this thesis we studied DNA copy number variation in human – how CNVs emerge and how they are inherited from parents to offspring. We also analysed CNVs in the context of few different diseases.
By using DNA microarrays we first aimed to determine if CNVs are associated with mental retardation (MR). For this we studied not only index cases with MR but larger nuclear families, where we discovered several already MR-associated CNVs and also a few novel CNV regions that are possibly associated with predisposition to MR.
Similar study was conducted in couples and females suffering from recurrent miscarriage. By comparing CNVs and their frequencies in the latter group to these of healthy mothers, we discovered a multi-copy duplication at 5p13.3 that disrupts PDZD2 and GOLPH3 genes and significantly increases maternal risk for pregnancy complications.
In the last part of this thesis we studied how CNVs are inherited in Estonian nuclear families (22 trios and 12 families with multiple siblings) and in HapMap Yoruban trios. We determined that deletion-carrying chromosomal regions were observed in the offspring slightly less frequently than expected by random Mendelian inheritance. By analysing duplication-carrying chromosomal regions in these families, we discovered that in two-thirds of such regions the duplicated copies of the underlying DNA sequence were not exactly identical but somewhat different, allowing us to define alternative allelic copies within these copy number gain-carrying chromosomal regions and demonstrating extensive and to-date unmeasured allelic variability in multi-copy CNV regions of the human genome
Systematic evaluation of NIPT aneuploidy detection software tools with clinically validated NIPT samples
No full textNon-invasive prenatal testing (NIPT) is a powerful screening method for fetal aneuploidy detection, relying on laboratory and computational analysis of cell-free DNA. Although several published computational NIPT analysis tools are available, no prior comprehensive, head-to-head accuracy comparison of the various tools has been published. Here, we compared the outcome accuracies obtained for clinically validated samples with five commonly used computational NIPT aneuploidy analysis tools (WisecondorX, NIPTeR, NIPTmer, RAPIDR, and GIPseq) across various sequencing depths (coverage) and fetal DNA fractions. The sample set included cases of fetal trisomy 21 (Down syndrome), trisomy 18 (Edwards syndrome), and trisomy 13 (Patau syndrome). We determined that all of the compared tools were considerably affected by lower sequencing depths, such that increasing proportions of undetected trisomy cases (false negatives) were observed as the sequencing depth decreased. We summarised our benchmarking results and highlighted the advantages and disadvantages of each computational NIPT software. To conclude, trisomy detection for lower coverage NIPT samples (e.g. 2.5M reads per sample) is technically possible but can, with some NIPT tools, produce troubling rates of inaccurate trisomy detection, especially in low-FF samples.sponsorship: This work was supported by the Ettevotluse Arendamise Sihtasutus [EU48695 to AS, EU53935 to AS and PPalta] (PP, HT, AS, KK, PPalta); and by the Eesti Teadusagentuur [PRG1076 to AS] (PP, HT, AS, KK, PPalta); and Horizon 2020 Framework Programme [EU952516 to AS] (PP, HT, AS, KK, PPalta). The funders had & nbsp;no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. (Ettevotluse Arendamise Sihtasutus|EU48695, Ettevotluse Arendamise Sihtasutus|EU53935, Eesti Teadusagentuur|PRG1076, Horizon 2020 Framework Programme|EU952516)status: Publishe
Denitrification in urban brownfield wetlands
No full textWetland processes are highly spatially and temporally heterogeneous, and managers lack models relating important wetland processes to specific combinations of biological communities, flooding, and soils. Wetlands in urban settings, while having the potential to deliver ecosystem services (nutrient removal) to urban areas, pose a particular challenge in linking ecosystem processes with their environmental drivers, because urban wetlands have been little studied, and each urban system has its own unique set of altered conditions. These issues are especially true of wetlands that develop on brownfield sites, on highly modified soil materials. My research questions were the following: (1) Where and when do the highest rates of NO3- removal occur in urban brownfield wetlands, and what are the spatial and temporal dimensions of these high rates?; (2) What are the environmental drivers of denitrification rates in urban brownfield wetlands?; and (3) How can the spatial and temporal dimensions of high NO3- removal rates be modeled and predicted to aid in restoration and management at the watershed scale? I utilized a combination of lab- and field-based studies to construct models designed to isolate and explain the relationship between environmental variables and soil denitrification rates in urban wetland environments. Whole-wetland denitrification potential was estimated through spatial interpolation of the variables mediating the highest rates of denitrification at the scale of a couple square meters. I also measured components of the nitrogen and hydrologic cycle in semi-permanently flooded wetlands to construct ecosystem budgets to estimate the role of denitrification in removing NO3- in these systems. My research shows that brownfield wetlands in northern New Jersey support active populations of denitrifying bacteria and are potential sinks for NO3- in urban landscapes. Rates of NO3- consumption in the soils equaled or exceeded the rate of NO3- loading, at least from the atmosphere. Soil structure and texture, water table levels, and landscape position appeared to be primary determinants of whether a brownfield soils have this ability. Modifications to hydrology that promote (1) endogenous NO3- production, particularly in low-oxygen waterlogged areas, and (2) contact between stormwater and soils with high macroporosity may augment levels of NO3- removal from soil.Ph. D.Includes bibliographical referencesby Monica Marie Palt
Notocomplana palta (Marcus 1954) Faubel 1983
No full textNotocomplana palta (Marcus 1954) Faubel 1983 Synonym: Notoplana palta Marcus 1954 Material examined. Specimens of Notocomplana palta were provided by the Swedish Museum of Natural History, Stockholm, Sweden (SMNH). We designate as lectotype the specimen SMNH 110097- 110101 from the series of animals studied by Marcus (1954). New specimens were collected in 2006–2009 at different localities along the Patagonian coast; these are housed in the Invertebrate Zoology collection at the MLP (Table 1, Figure 1). Description. Live specimens are light to dark orange-colored (Figure 5 B). Central zone where pharynx is placed is darker. Body ovate and with ruffled edges. Studied specimens measure 10–20 mm long and 5–9 mm wide. Tentacles absent. Eyes placed in medial region, associated to brain zone. Brain eyes rounded, small, numbering 35 to 40 on each side. Tentacular eyes posterior to brain eyes, larger, and approximately ten on each side. Very small eyes in parenchyma ventrally and anteriorly to brain (Figure 5 A). Ventral and dorsal epidermis ciliated. Ventral cilia longer. Rhabdites on both surfaces, more abundant on ventral epidermis. Basal membrane thick, the ventral one being thinner than the height of the epithelium; dorsal basal membrane thickness equal to the epithelium height, folded in some of the thin sections. Body-wall musculature on dorsal surface consisting of an external thin circular layer, a diagonal middle layer (the thickest) and a longitudinal internal layer; a second internal circular layer is crossed by dorso-ventral muscles. On the ventral surface of the body, musculature is longitudinal, followed internally by a thin circular layer; the latter is packed inwards. Pharynx ruffled (approximately nine folds), placed in front of mid-length of body. Epithelium lining the pharynx cavity is ciliated. Mouth slightly behind mid-length of pharynx (Figure 5 C). Testicles ventral and ovaries dorsal, distributed throughout body length, including the region in front of the brain. Deferent ducts (one pair) thickenning and twisting before joining seminal vesicle. Seminal vesicle strongly muscular, rounded to oval. Ejaculatory duct long, thin and muscular. A short portion of ejaculatory duct projects into prostatic vesicle, which shows a tall and folded epithelium. Lumen of prostatic vesicle continuous with the lumen of penis papilla (Figure 4). Penis papilla conical, located in a male atrium and opening into a male pore (Figure 5 E, H–I). Female gonopore posterior to male pore (Figure 4). Muscular vagina inlaid with ciliate epithelium; shape sigmoidal, and lying perpendicular to surface in its first part, then running anteriorly, and later turning posteriorly. In female specimens, the portion reaching anteriorly carries abundant cement glands that open into that part of the vagina (Figures 4, 5 D, F). Uteri loaded with ovocites and continuing into two uterine ducts that join to form a single ciliate duct. The latter joins the proximal part of vagina at its ventral surface. Lang´s vesicle channel present after junction of uterus and vagina. This channel shows constrictions that give it a beaded appearance; it opens into Lang’s vesicle. At the junction of vesicle and duct there is a sphincter. Vesicle rounded to ovate, lined with a tall vacuolar epithelium, with scarce muscular fibres. Sperms were observed within it (Figures 4, 5 D, G). Remarks. This species was originally described by Marcus (1954) from the fjord region in southern Chile. The Patagonian specimens clearly agree with the original description of the species (Marcus, 1954) and with the material he had available for study (SMNH 110097- 110101). Bulnes (2009) recently mentioned Notocomplana palta in an area close to the type locality but the description given by this author does not agree with the original desciption (Marcus, 1954). Bulnes (2009) described small tentacles, a thin and short penis papilla, and a four-chambered seminal vesicle. These structures were not mentioned in the original description of this species (Marcus, 1954) nor are they present in the material studied by him and housed in the SMNH. Therefore, they can not be considered part of this taxon. The material housed in the SMNH was not designated originally as type material. To avoid future misidentifications of Notocomplana palta, the specimen described and illustrated by Marcus in the original work (Marcus 1954; 65; figs. 99, 101) and from the type locality (as recorded on the original label St. M 21) is designated as the lectotype under Article 74 of the fourth edition of ICZN (1999). The presence of this species along the Patagonian coast and southern Chile suggests a Magellanic distribution similar to that of other species of invertebrates (e.g. Boschi 2000, Balech & Ehrlich 2008).Published as part of Brusa, Francisco & Damborenea, Cristina, 2011, Polycladida Acotylea from Patagonia. Redescription of Crassiplana albatrossi (Pseudostylochidae), lectotype designation and first record of Notocomplana palta (Notoplanidae), pp. 29-38 in Zootaxa 2903 on pages 34-36, DOI: 10.5281/zenodo.20113
Author Correction: FinnGen provides genetic insights from a well-phenotyped isolated population
No full textSuunatud ja ülegenoomsel sekveneerimisel põhinevate mitteinvasiivsete sünnieelsete testide arvutusmeetodite ja töövoogude väljatöötamine
No full textVäitekirja elektrooniline versioon ei sisalda publikatsiooneLoote sõeluuring võimaldab avastada lootel esinevaid arenguhäireid ja sagedasemaid kromosoomhaiguseid, nagu näiteks Down’i, Edwards’i ja Patau sündroom. Varajane teave lootel esineva kromosoomhaiguse kohta võimaldab langetada informeeritud otsust raseduse jätkamise osas ning aitab tulevasi vanemaid paremini ette valmistada.
Tavapärane loote sõeluuring sisaldab loote ultraheli uuringut ja vereseerumi analüüsi, mille abil tuvastatakse enamik kromosoomhaigusega loodetest. Lõpliku diagnoosi saamiseks suunatakse kõrge riski saanud patsient edasi invasiivsele protseduurile. Eelnimetatud sõeluuringute puuduseks on arvestatav valepositiivsete hulk, mistõttu enamik positiivse testitulemuse saanud patsientidest kannab täiesti tervet loodet. Sõeluuringule järgnev invasiivne protseduur on neil juhtudel ebavajalik, põhjustab rasedatele asjatut stressi ning sellega võib kaasneda suurenenud oht raseduse katkemiseks.
Antud doktoritöö keskseks teemaks on mitte-invasiivne sünnieelne testimine (NIPT), mis põhineb ema veres leiduva loote päritolu rakuvaba DNA analüüsil. Võrreldes eelmainitud traditsionaalsete sõeluuringu meetoditega, on NIPT oluliselt sensitiivsem ja spetsiifilisem sagedamini esinevate kromosoomihäirete avastamiseks.
Doktoritöö raames arendati välja TAC-seq põhine analüüsi töövoog, mida rakendati 21. kromosoom trisoomia tuvastamiseks. Lisaks töötati välja NIPT analüüsiraamistik, mis kasutab erinevaid masinõppe metoodikaid loote trisoomia määramiseks rakuvaba DNA-st. Niisamuti viidi Eesti rasedate kohordil läbi NIPT metoodika validatsiooni uuring, milles rakendati ülegenoomsel sekveneerimisel põhinevat töövoogu sagedamate loote kromosoomihäirete määramiseks.
Üldiselt on nii suunatud kui ka ülegenoomsel NIPT meetoditel muutnud rasedate sõeluuring varasemast veel täpsemaks. Kui suunatud sekveneerimise suureks eeliseks on kulutõhusus, siis ülegenoomne lähenemine tuvastab valimatult kõikvõimalikke geneetilisi aberratsioone üle kogu genoomi.Fetal screening allows to detect congenital anomalies and more frequent chromosomal abnormalities, such as Down, Edwards and Patau syndrome. Early information about a fetus’s possible health problem allows to make an informed decision about the continuation of the pregnancy and better prepare the future parents.
Conventional screening includes an ultrasound and blood serum analysis by way of which most of the fetal chromosomal abnormalities are detected. For a final diagnosis, the patients who are deemed to have a high risk for fetal chromosomal aberrations are referred to an invasive procedure. The disadvantage of the aforementioned screening method is a considerable number of false positive results, which is why most of the patients who receive a positive result are actually carrying a fully healthy fetus. The invasive procedure that follows the screening is unnecessary for those patients, causes them undue stress and this may also lead to a higher risk of miscarriage.
The focal point of this doctoral thesis is non-invasive prenatal testing (NIPT), which is based on the analysis of cell-free DNA (cfDNA) of fetal origin that is found in maternal blood. In comparison to the above-mentioned conventional screening methods, NIPT is considerably more sensitive and specific for detecting the most common chromosomal abnormalities.
In the framework of the thesis, TAC-seq based analysis workflow was developed and used to detect chromosome 21 trisomy. In addition, NIPT analysis framework, which uses different machine learning methods, was developed for determining fetal trisomies from cfDNA sample. Also, a validation study of NIPT was carried out on pregnant women in Estonian cohort using a whole-genome sequencing based workflow.
In general, both targeted and whole-genome sequencing based NIPT methods have made prenatal screening of fetal aneuplodies even more accurate than before. While cost-effectiveness is a major advantage of the targeted sequencing based approach, the whole-genome sequencing based NIPT possibly detects all kinds of genetic aberrations across the genome.https://www.ester.ee/record=b549777
Screening of 20 patients with X-linked mental retardation using chromosome X-specific array-MAPH
No full textCopyright © 2007 Elsevier Masson SAS All rights reserved.The rapid advancement of high-resolution DNA copy number assessment methods revealed the significant contribution of submicroscopic genetic imbalances to abnormal phenotypes, including mental retardation. In order to detect submicroscopic genetic imbalances, we have screened 20 families with X-linked mental retardation (XLMR) using a chromosome X-specific array-MAPH platform with median resolution of 238 kb. Among the 20 families, 18 were experimental, as they were not previously screened with any microarray method, and two were blind controls with known aberrations, as they were previously screened by array-CGH. This study presents the first clinical application of chromosome X-specific array-MAPH methodology. The screening of 20 affected males from 20 unrelated XLMR families resulted in the detection of an unknown deletion, spanning a region of 7-23 kb. Family studies and population screening demonstrated that the detected deletion is an unknown rare copy number variant. One of the control samples, carrying approximately 6-Mb duplication was correctly identified, moreover it was found to be interrupted by a previously unknown 19 kb region of normal copy number. The second control 50 kb deletion was not identified, as this particular region was not covered by array-MAPH probes. This study demonstrates that the chromosome X-specific array-MAPH platform is a valuable tool for screening patients with XLMR, or other X-linked disorders, and emerges the need for introducing new high-resolution screening methods for the detection of genetic imbalances. (C) 2007 Elsevier Masson SAS. All fights reserved.Ludmila Kousoulidou, Sven Parkel, Olga Zilina, Priit Palta, Helen Puusepp, Maido Remm, Gillian Turner, Jackie Boyle, Hans van Bokhoven, Arjan de Brouwer, Hilde Van Esch, Guy Froyen, Hans-Hilger Ropers, Jamel Chelly, Claude Moraine, Jozef Gecz, Ants Kurg and Philippos C. Patsalishttp://www.elsevier.com/wps/find/journaldescription.cws_home/705239/description#descriptio
Computational framework for targeted high-coverage sequencing based NIPT
No full textNon-invasive prenatal testing (NIPT) enables accurate detection of fetal chromosomal trisomies. The majority of publicly available computational methods for sequencing-based NIPT analyses rely on low-coverage whole-genome sequencing (WGS) data and are not applicable for targeted high-coverage sequencing data from cell-free DNA samples. Here, we present a novel computational framework for a targeted high-coverage sequencing-based NIPT analysis. The developed framework uses a hidden Markov model (HMM) in conjunction with a supplemental machine learning model, such as decision tree (DT) or support vector machine (SVM), to detect fetal trisomy and parental origin of additional fetal chromosomes. These models were developed using simulated datasets covering a wide range of biologically relevant scenarios with various chromosomal quantities, parental origins of extra chromosomes, fetal DNA fractions, and sequencing read depths. Developed models were tested on simulated and experimental targeted sequencing datasets. Consequently, we determined the functional feasibility and limitations of each proposed approach and demonstrated that read count-based HMM achieved the best overall classification accuracy of 0.89 for detecting fetal euploidies and trisomies on simulated dataset. Furthermore, we show that by using the DT and SVM on the HMM classification results, it was possible to increase the final trisomy classification accuracy to 0.98 and 0.99, respectively. We demonstrate that read count and allelic ratio-based models can achieve a high accuracy (up to 0.98) for detecting fetal trisomy even if the fetal fraction is as low as 2%. Currently, existing commercial NIPT analysis requires at least 4% of fetal fraction, which can be possibly a challenge in case of early gestational age (35 kg/m2). More accurate detection can be achieved at higher sequencing depth using HMM in conjunction with supplemental models, which significantly improve the trisomy detection especially in borderline scenarios (e.g., very low fetal fraction) and enables to perform NIPT even earlier than 10 weeks of pregnancy.</div
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