An accurate and rapid gender determination assay in single cells by the capillary polymerase chain reaction method

Purpose: In preimplantation genetic diagnosis (PGD), a rapid and accurate assay has been required. We have therefore developed a capillary palymerase chain reaction (PCR) method using rapid thermal cycling programs to determine the gender of single amniocytes. Methods: Single amniocytes from each amniotic fluid sample were isolated by micromanipulation and their gender was determined by a multiplex PCR assay in a capillary tube, using primers that amplify a 308-bp DXZI and a 154-bp DYZI repeat sequence on the X and Y chromosomes, respectively. Results: All four thermal cycling programs, which took 180, 150, 120, and 90 min, were 100% accurate in diagnosing the gender of single amniocytes. No DNA contamination was observed in any samples. Conclusions: The multiplex PCR assay was rapid and accurate in diagnosing gender in single cells and may be clinically applicable in PGD.