Cíl studie: Zhodnocení kaskádového vyšetření metodami kvantitativní fluorescenční PCR (QF-PCR – quantitative fluorescence PCR) a SNP array (single nucleotide polymorphism array) při identifikaci chromozomálních abnormalit u potracených plodů. Soubor a metodika: V průběhu let 2018–2020 bylo v laboratořích Gennetu zpracováno 1 094 vzorků potracených plodů. Standardní schéma kaskádového vyšetření zahrnovalo screening základních aneuploidií metodou QF-PCR setem Omnibor (STR markery 13, 18, 21, X a Y), setem SAB-nadstavba I (STR markery 2, 7, 15, 16, 22), setem SAB-nadstavba II (od listopadu 2019, STR markery 4, 6, 14) a následně vyšetření metodou SNP array (Illumina). U všech vzorků bylo provedeno ověření/vyloučení maternální kontaminace. Výsledky: Po vyloučení maternální kontaminace (32 % vzorků) bylo metodou QF-PCR vyšetřeno 742 vzorků, 469 (63,2 %) z nich mělo negativní nález a 273 (36,8 %) patologický nález. Vzorky 469 plodů s negativním QF-PCR nálezem byly následně vyšetřeny metodou SNP array. Normální ženský/mužský profil byl potvrzen u 402 plodů (85,7 %), chromozomální aneuploidie a velké chromozomální aberace (delece/duplikace > 10 Mb) u 51 vzorků (10,9 %). U 16 (3,4 %) vyšetřených plodů byla nalezena mikrodelece nebo mikroduplikace, ve dvou případech se jednalo o patogenní mikrodelece a ve 14 o variantu nejasného významu bez přímé souvislosti s abortem. Byl potvrzen statisticky významný vyšší záchyt chromozomálních aberací u plodů žen s věkem > 35 let. Mezi skupinami vyšetřených abortů u gravidit po spontánní koncepci a po in vitro fertilizaci nebyl prokázán statisticky významný rozdíl. Závěr: Kaskádovým vyšetřením metodami QF-PCR a SNP array byla objasněna genetická příčina u 43 % všech vyšetřených abortů. Záchyt chromozomálních abnormalit u časných abortů v I. trimestru byl 50,4 %.
Objective: The evaluation of quantitative fluorescence PCR (QF-PCR) and single nucleotide polymorphism array (SNP array) analysis for the identification of chromosomal abnormalities in products of conception (POC). Materials and methods: A total of 1,094 POC samples were processed at Gennet in the years 2018–2020. Chromosomal aneuploidies were tested by QF-PCR using a Omnibor set (STR markers 13, 18, 21, X a Y), SAB-I set (STR markers 2, 7, 15, 16, 22), SAB-II set (from November 2019, STR markers 4, 6, 14) followed by SNP array analysis (Illumina) on samples with a negative QF-PCR result. All POC samples were tested for maternal contamination. Results: After exclusion of maternal contamination (32% samples) the total number of 742 POC samples were tested by QF-PCR. Chromosomal aneuploidies were found in 273 POC samples (36.8%). Then, 469 QF-PCR negative POC samples were tested by SNP array analysis. Normal female/male profile was confirmed in 402 samples (85.7%) and chromosomal aneuploidies and chromosomal aberrations (deletion/duplication > 10 Mb) in 51 samples (10.9%). Microdeletion/microduplication was found in 16 POC samples (3.4%), two were classified as pathogenic variants and 14 as variants of unknown significance. In a group of women > 35 years of age, statistically significant increase of the chromosomal abnormalities was confirmed. No statistically significant difference between the in vitro fertilization group and the group of spontaneous conception was found. Conclusion: The application of the molecular work-up based on the stepwise use of QF-PCR and SNP array clarifies the cause of the abortion in 43% POC samples. The overall detection rate in the I. trimester was 50.4%.
OBJECTIVE: To present methodical approach of preimplantation genetic diagnosis (PGD) as an option for an unaffected pregnancy in reproductive-age couples who have a genetic risk of the X-linked dominant peripheral neuropathy Charcot-Marie-Tooth type 1 disease. PATIENTS AND METHODS: We performed PGD of X-linked Charcot-Marie-Tooth type 1 disease using haplotyping/indirect linkage analysis, when during analysis we reach to exclude embryos that carry a high-risk haplotype linked to the causal mutation p.Leu9Phe in the GJB1 gene. RESULTS: Within the PGD cycle, we examined 4 blastomeres biopsied from cleavage-stage embryos and recommended 3 embryos for transfer. Two embryos were implanted into the uterus; however, it resulted in a singleton pregnancy with a male descendant. Three years later, the couple returned again with spontaneous gravidity. A chorionic biopsy examination of this gravidity ascertained the female sex and a pericentric inversion of chromosome 5 in 70% of the cultivated foetal cells. CONCLUSION: Using indirect linkage analysis, PGD may help to identify genetic X-linked defects within embryos during screening, thereby circumventing the potential problems with abortion.
- MeSH
- Charcotova-Marieova-Toothova nemoc diagnóza genetika MeSH
- genetická vazba MeSH
- genetické markery MeSH
- genetické testování metody MeSH
- haplotypy MeSH
- konexiny genetika MeSH
- lidé MeSH
- mutace MeSH
- preimplantační diagnóza metody MeSH
- těhotenství MeSH
- Check Tag
- lidé MeSH
- těhotenství MeSH
- ženské pohlaví MeSH
- Publikační typ
- časopisecké články MeSH
- kazuistiky MeSH
Approximately 20 cases of genome-wide uniparental disomy or diploidy (GWUPD) as mosaicism have previously been reported. We present the case of an 11-year-old deaf girl with a paternal uniparental diploidy or isodisomy with a genome-wide loss of heterozygosity (LOH). The patient was originally tested for non-syndromic deafness, and the novel variant p.V234I in the ESRRB gene was found in a homozygous state. Our female proband is the seventh patient diagnosed with GWUPD at a later age and is probably the least affected of the seven, as she has not yet presented any malignancy. Most, if not all, reported patients with GWUPD whose clinical details have been published have developed malignancy, and some of those patient developed malignancy several times. Therefore, our patient has a high risk of malignancy and is carefully monitored by a specific outpatient pediatric oncology program. This observation seems to be novel and unique in a GWUPD patient. Our study is also unique as it not only provides very detailed documentation of the genomic situations of various tissues but also reports differences in the mosaic ratios between the blood and saliva, as well as a normal biparental allelic situation in the skin and biliary duct. Additionally, we were able to demonstrate that the mosaic ratio in the blood remained stable even after 3 years and has not changed over a longer period.
- MeSH
- celogenomová asociační studie MeSH
- diploidie * MeSH
- dítě MeSH
- exprese genu MeSH
- hluchota diagnóza genetika patofyziologie MeSH
- lidé MeSH
- mozaicismus * MeSH
- mutace * MeSH
- nestabilita genomu MeSH
- receptory pro estrogeny genetika MeSH
- rodokmen MeSH
- sekvence nukleotidů MeSH
- sekvenční analýza DNA MeSH
- uniparentální disomie * MeSH
- ztráta heterozygozity MeSH
- Check Tag
- dítě MeSH
- lidé MeSH
- ženské pohlaví MeSH
- Publikační typ
- časopisecké články MeSH
- kazuistiky MeSH
- MeSH
- dospělí MeSH
- fertilizace in vitro MeSH
- genetická vazba MeSH
- genetické nemoci vázané na chromozom X * diagnóza genetika prevence a kontrola MeSH
- haplotypy MeSH
- kohortové studie MeSH
- lidé MeSH
- mikrosatelitní repetice MeSH
- mutace MeSH
- preimplantační diagnóza * metody MeSH
- přenos embrya MeSH
- reprodukovatelnost výsledků MeSH
- retrospektivní studie MeSH
- techniky amplifikace nukleových kyselin MeSH
- těhotenství MeSH
- úhrn těhotenství na počet žen v reprodukčním věku MeSH
- Check Tag
- dospělí MeSH
- lidé MeSH
- mužské pohlaví MeSH
- těhotenství MeSH
- ženské pohlaví MeSH
- Geografické názvy
- Česká republika MeSH
Charcot-Marie-Tooth (CMT) disease is a heterogeneous disorder of the peripheral nervous system that collectively affects approximately 1 in 2,500 individuals, thus making it the most common inherited neurologic disorder. X-linked inheritance may account for 10-20 % of CMT neuropathy. We report a Czech family with a 30-year-old woman affected by CMT since the age of 10 years, originally as an isolated case. Nerve conduction study (NCS) showed demyelinating neuropathy, and DNA testing revealed a novel heterozygous gap junction beta-1 protein (GJB1) mutation c.784_786delTA. The same mutation, but surprisingly in heterozygous state, was subsequently found in her subjectively healthy father and later also in one of her sisters but not in her two other sisters. NCS showed intermediate type of motor and sensory neuropathy in these two females manifesting heterozygotes and normal results in the other healthy sisters and one brother, all without the c.784_786delTA mutation. The father has a phenotype milder than his daughter and has only subclinical signs of CMT. The index female patient had normal karyotype 46, XX, and normal FISH for centromeric X chromosome. We concluded that the proband's father is a heterozygote due to the somatic mosaicism for the GJB1 mutation in his leukocytes (detected by DNA sequencing) and also in his germ cells as confirmed by the unexpectedly different genotypes in his four daughters. Quantitative analysis revealed a mutated signal in 25:75 allele proportion of mutated to healthy allele in the mosaic father. This study has important consequences for genetic counseling and prognosis in CMTX1 families.
- MeSH
- Charcotova-Marieova-Toothova nemoc diagnóza genetika MeSH
- dospělí MeSH
- geny vázané na chromozom X MeSH
- heterozygot * MeSH
- konexiny genetika MeSH
- lidé středního věku MeSH
- lidé MeSH
- mozaicismus * MeSH
- periferní nervy patofyziologie MeSH
- Check Tag
- dospělí MeSH
- lidé středního věku MeSH
- lidé MeSH
- mužské pohlaví MeSH
- ženské pohlaví MeSH
- Publikační typ
- časopisecké články MeSH
- kazuistiky MeSH
- Research Support, N.I.H., Extramural MeSH
- Geografické názvy
- Česká republika MeSH
- Publikační typ
- abstrakt z konference MeSH
Non-Governmental Organization Archaia (http://www.archaia.cz) carried out the rescue archaeological research at Knezeves near Prague in 1998. Most of dating objects in Knezeves come from the period of Late and Final Bronze Age. The approximately 3,000 years old set, which included 11 human remains from three settlement features, was collected for the study. First, gender was determined according to anthropological characteristics. Ancient DNA from bones was extracted by the phenol-chloroform procedure and N-phenacetylthiazolum bromide reagent. Polymerase chain reaction amplification of AMEL XY, part of amelogenin gene, with subsequent polyacrylamide gel electrophoresis and Short Tandem Repeats analysis followed. DNA profiles of skeletal remains were obtained by the fragmentation analysis of autosomal short tandem repeat markers. Genetic profiles showed us whether individuals from Knezeves were in mutual relationship (parent - descendant). The congruence of results in sex determination supported reliability of genetic methods, which are suitable for sex determination of fragmental and subadult skeletal remains.
- MeSH
- archeologie metody MeSH
- dějiny starověku MeSH
- DNA analýza MeSH
- kosti a kostní tkáň chemie MeSH
- lidé MeSH
- mikrosatelitní repetice MeSH
- polymerázová řetězová reakce metody MeSH
- sekvence nukleotidů MeSH
- sekvenční analýza DNA metody MeSH
- Check Tag
- dějiny starověku MeSH
- lidé MeSH
- mužské pohlaví MeSH
- ženské pohlaví MeSH
- Publikační typ
- historické články MeSH
- publikace stažené z tisku MeSH
The settlement at Kněževes is situated near Prague, and the non-profit company Archaia carried out a rescue archaeological research there in 1998. A set of skeletal remains was collected from three settlement features dated in the Late Bronze Age. The skeletons were subjected to anthropological and genetic analyses for sex determination. First, in the genetic analysis recent DNA from the femur was used to compare isolation methods. We tested four different DNA extraction protocols: phenol-chloroform method, Dextran Blue mediated extraction method, and two isolation protocols using the DNA IQTM System, and the QIAamp DNA Blood Mini Kit, respectively. Skeletal remains from Kněževes were subjected to the selected extraction method and subsequently to PCR (Polymerase Chain Reaction) reaction for sex determination. The amplified aDNA (ancient DNA) was separated using the polyacrylamide electrophoresis. After comparison of genetic and anthropological methods, aDNA analysis confirmed that it is reliable for sex determination of subadult skeletal remains with undeveloped secondary sex characteristics.
