title: Refined quantitative fluorescent PCR of Y-chromosome DNA sequences mosaics in Turner's syndrome patients--alternative to real-time PCR.
abstract: BACKGROUND
  Real-time polymerase chain reactions (PCRs) are the most frequently used techniques for gonosomal mosaics quantification. The primary aim of this work is to assess and optimize the refined technique of quantitative fluorescent polymerase chain reaction (RQF PCR) in the quantification of Y-chromosome sequences in gonosomal mosaics. The method was applied to the analysis of Y-chromosome sequences (amelogenin gene, AMELX/Y-loci) in peripheral lymphocytes and gonadal tissues in Y-positive Turner's syndrome (TS) patients.
  
  
  METHODS
  RQF PCR was used for molecular quantification, and fluorescent in situ hybridization (FISH) technique was used for comparison.
  
  
  RESULTS
  Based on a formulated calibration curve, DNA mosaics from six Y-positive patients and gonads from one patient were deducted. For calculation of rare mosaics, it is possible to take advantage of a new empirical formula. FISH results were comparable to RQF PCR.
  
  
  CONCLUSIONS
  The sensitivity of RQF PCR brings significant progress in the analysis of gonosomal aberrations. RQF PCR also finds applications in prenatal diagnostics of maternal contaminations of amniotic fluid and foetal DNA in maternal blood and analysis of chimerism in patients after bone marrow transplantation. The method is very convenient for determining the number of testis-specific protein, Y-linked (TSPY) gene repetitions.
authors.affiliation: Department of Clinical Genetics and Foetal Medicine, Palacky University, University Hospital Olomouc, I.P Pavlova 6, 775 20 Olomouc, Czech Republic, vodickar@finol.cz
authors.forename: Radek
authors.initials: R
authors.lastname: Vodicka
authors.forename: Radek
authors.initials: R
authors.lastname: Vrtel
authors.forename: Ondrej
authors.initials: O
authors.lastname: Scheinost
authors.forename: Jirina
authors.initials: J
authors.lastname: Zapletalova
authors.forename: Ladislav
authors.initials: L
authors.lastname: Dusek
authors.forename: Geierova
authors.initials: G
authors.lastname: Marie
authors.forename: Jiri
authors.initials: J
authors.lastname: Santavy
mesh_terms: D002138:Calibration; D002869:Chromosome Aberrations; D041321:Chromosomes, Human, X; D041322:Chromosomes, Human, Y; D004247:DNA / Q000737:chemistry; D019075:Electrophoresis, Capillary; D006801:Humans; D017404:In Situ Hybridization, Fluorescence; D016014:Linear Models; D008297:Male; D008856:Microscopy, Fluorescence; D015233:Models, Statistical; D009030:Mosaicism; D016133:Polymerase Chain Reaction / Q000379:methods*; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods*; D014424:Turner Syndrome / Q000235:genetics*
chemical_list: D004247:DNA