Non-invasive fetal sex diagnosis in plasma of early weeks pregnants using droplet digital PCR

Elisabetta D’Aversa, Giulia Breveglieri, Patrizia Pellegatti, Giovanni Guerra, Roberto Gambari, Monica Borgatti

Fetal sex determination is useful for families at risk of X-linked disorders, such as Duchenne muscular dystrophy, adrenal hypoplasia, hemophilia. At first, this could be obtained through invasive procedures such as amniocentesis and chorionic villus sampling, having a 1% risk of miscarriage. Since the discovery of cell-free fetal DNA (cffDNA) in maternal plasma, noninvasive prenatal testing permits the early diagnosis of fetal sex through analysis of cffDNA. However, the low amount of cffDNA relative to circulating maternal DNA requires highly sensitive molecular techniques in order to perform noninvasive prenatal diagnosis. In this context we employed droplet digital PCR (ddPCR) in order to evaluate the earliest possible fetal sex determination from circulating DNA extracted from plasma of pregnant women at different gestational ages. We identified the fetal sex on cffDNA extracted from 29 maternal plasma samples at early gestational ages, several of them not suitable for qPCR determination, using ddPCR designed for SRY gene target. All maternal plasma samples were determined correctly for SRY gene target using ddPCR even at very early gestational age (prior to 7 weeks). The ddPCR is a robust, efficient and reliable technology for the earliest possible fetal sex determination from maternal plasma.