title: Development of novel noninvasive prenatal testing protocol for whole autosomal recessive disease using picodroplet digital PCR.
abstract: We developed a protocol of noninvasive prenatal testing (NIPT), employing a higher-resolution picodroplet digital PCR, to detect genetic imbalance in maternal plasma DNA (mpDNA) caused by cell-free fetal DNA (cffDNA). In the present study, this approach was applied to four families with autosomal recessive (AR) congenital sensorineural hearing loss. First, a fraction of the fetal DNA in mpDNA was calculated. Then, we made artificial DNA mixtures (positive and negative controls) to simulate mpDNA containing the fraction of cffDNA with or without mutations. Next, a fraction of mutant cluster signals over the total signals was measured from mpDNA, positive controls, and negative controls. We determined whether fetal DNA carried any paternal or maternal mutations by calculating and comparing the sum of the log-likelihood of the study samples. Of the four families, we made a successful prediction of the complete fetal genotype in two cases where a distinct cluster was identified for each genotype and the fraction of cffDNA in mpDNA was at least 6.4%. Genotyping of only paternal mutation was possible in one of the other two families. This is the first NIPT protocol potentially applicable to any AR monogenic disease with various genotypes, including point mutations.
authors.affiliation: Department of Otorhinolaryngology-Head and Neck Surgery, Chung-Ang University College of Medicine, 102 Heukseok-ro, Dongjak-gu, Seoul, 06973, Republic of Korea.
authors.forename: Mun Young
authors.initials: MY
authors.lastname: Chang
authors.affiliation: Department of Otorhinolaryngology, Seoul National University Hospital, Seoul national University College of Medicine, 101 Daehak-ro, Jongno-gu, Seoul 03080, Republic of Korea.
authors.forename: Ah Reum
authors.initials: AR
authors.lastname: Kim
authors.affiliation: Department of Otorhinolaryngology, Seoul National University Bundang Hospital, 82 Gumi-ro 173 beon-gil, Bundang-gu, Seongnam, 13620, Republic of Korea.
authors.forename: Min Young
authors.initials: MY
authors.lastname: Kim
authors.affiliation: LAS Inc., 16 Arayuk-ro, Gimpo, 10136, Republic of Korea.
authors.forename: Soyoung
authors.initials: S
authors.lastname: Kim
authors.affiliation: Bio-Medical Science Co., Ltd., BMS Bldg., 22 Yeoksam-ro 7-gil, Gangnam-gu, Seoul 06244, Republic of Korea.
authors.forename: Jinsun
authors.initials: J
authors.lastname: Yoon
authors.affiliation: Department of Otorhinolaryngology, Seoul National University Bundang Hospital, 82 Gumi-ro 173 beon-gil, Bundang-gu, Seongnam, 13620, Republic of Korea.
authors.forename: Jae Joon
authors.initials: JJ
authors.lastname: Han
authors.affiliation: Medical Research Collaborating Center, Seoul National University Bundang Hospital, 82 Gumi-ro 173 beon-gil, Bundang-gu, Seongnam, 13620, Republic of Korea.
authors.forename: Soyeon
authors.initials: S
authors.lastname: Ahn
authors.affiliation: Department of Biology and Research Institute of Basic Sciences, College of Natural Sciences, Sungshin Women's University, Dongseon-dong 3(sam)-ga, Seongbuk-gu, Seoul, 01133, Republic of Korea.
authors.forename: Changsoo
authors.initials: C
authors.lastname: Kang
authors.affiliation: Department of Otorhinolaryngology, Seoul National University Bundang Hospital, 82 Gumi-ro 173 beon-gil, Bundang-gu, Seongnam, 13620, Republic of Korea.|Wide River Institute of Immunology, Seoul National University College of Medicine, 101 Dabyeonbat-gil, Hwachon-myeon, Hongcheon, 25159, Republic of Korea.
authors.forename: Byung Yoon
authors.initials: BY
authors.lastname: Choi
mesh_terms: D001800:Blood Specimen Collection; D000072259:Connexin 26; D017630:Connexins / Q000235:genetics; D004247:DNA / Q000097:blood / Q000302:isolation & purification; D004252:DNA Mutational Analysis / Q000379:methods*; D016002:Discriminant Analysis; D005260:Female; D005315:Fetal Diseases / Q000175:diagnosis* / Q000235:genetics; D005331:Fetomaternal Transfusion / Q000097:blood / Q000235:genetics*; D005808:Genes, Recessive; D060005:Genotyping Techniques; D006319:Hearing Loss, Sensorineural / Q000175:diagnosis / Q000196:embryology / Q000235:genetics; D006801:Humans; D008297:Male; D026901:Membrane Transport Proteins / Q000235:genetics; D008832:Microchemistry / Q000379:methods*; D025202:Molecular Diagnostic Techniques; D016133:Polymerase Chain Reaction; D020641:Polymorphism, Single Nucleotide; D011247:Pregnancy; D011296:Prenatal Diagnosis / Q000379:methods*; D017422:Sequence Analysis, DNA; D000076988:Sulfate Transporters
chemical_list: D017630:Connexins; C000607042:GJB2 protein, human; D026901:Membrane Transport Proteins; C085609:SLC26A4 protein, human; D000076988:Sulfate Transporters; D000072259:Connexin 26; D004247:DNA