BACKGROUND: @entity1062 ( @entity1062 ) for monogenic diseases by use of PCR-based strategies requires precise quantification of mutant fetal alleles circulating in the maternal plasma. The study describes the development and validation of a novel assay termed circulating single-molecule amplification and resequencing technology (cSMART) for counting single allelic molecules in plasma. Here we demonstrate the suitability of cSMART for @entity1062 , with @entity2360 ( @entity2360 ) as proof of concept. METHODS: We used Sanger and whole-exome sequencing to identify familial @entity37824 ( @entity4430 , Cu(++) transporting, b polypeptide) gene mutations. For cSMART, single molecules were tagged with unique barcodes and circularized, and alleles were targeted and replicated by inverse PCR. The unique single allelic molecules were identified by sequencing and counted, and the percentage of mutant alleles in the original maternal plasma sample was used to determine fetal genotypes. RESULTS: Four families with @entity2360 pedigrees consented to the study. Using Sanger and whole-exome sequencing, we mapped the pathogenic @entity37824 mutations in each pedigree and confirmed the proband's original diagnosis of @entity2360 . After validation of cSMART with defined plasma models mimicking fetal inheritance of paternal, maternal, or both parental mutant alleles, we retrospectively showed in second pregnancies that the fetal genotypes assigned by invasive testing and @entity1062 were concordant. CONCLUSIONS: We developed a reliable and accurate @entity1062 assay that correctly diagnosed the fetal genotypes in 4 pregnancies at risk for @entity2360 . This novel technology has potential as a universal strategy for @entity1062 of other monogenic disorders, since it requires only knowledge of the parental pathogenic mutations.

Noninvasive Prenatal Testing for XXXX by Use of Circulating Single-Molecule Amplification and Resequencing Technology (cSMART).

@entity37824 :: ('540', 'Gene') :: ['ATP7B'] @entity4430 :: ('1769', 'Gene') :: ['ATPase'] @entity2360 :: ('MESH:D006527', 'Disease') :: ['Wilson Disease', 'WD'] @entity1062 :: ('MESH:D013736', 'Disease') :: ['Noninvasive prenatal testing', 'NIPT']
@entity2360 :: (MESH:D006527,Disease) :: ['Wilson Disease']
