Detection of a Paternally Inherited Fetal Mutation in Maternal Plasma by the Use of Automated Sequencing ANA BUSTAMANTE-ARAGONES, MARIA GARCIA-HOYOS, MARTA RODRIGUEZ DE ALBA, CRISTINA GONZALEZ-GONZALEZ, ISABEL LORDA-SANCHEZ, DAN DIEGO-ALVAREZ, M. JOSE TRUJILLO-TIEBAS, CARMEN AYUSO, AND CARMEN RAMOS Department of Genetics, Fundacion Jimenez Diaz, Madrid, Spain ABSTRACT: The discovery of circulating fetal DNA in maternal blood has been an encouraging step forward in the prenatal diagnostic field. It has opened up the possibility of development of a noninvasive method for the genetic analysis of the fetus. Many techniques have been applied to the study of this fetal DNA, but automated sequencing has been seldom used. The intention of this study was to use the automated sequencing technique for the detection of a paternally inherited fetal mutation in maternal plasma. Maternal plasma samples from a pregnant woman, whose husband had a mutation (Q134X) in the RP2 gene, which is lo- cated in the X-chromosome, were collected at two different gestational ages (10th and 19th week of gestation) in order to determine whether the paternally inherited fetal mutation could be detected by automated se- quencing. Restriction analysis was also performed to confirm the results. The fetal mutation was clearly detected in the maternal plasma by the use of automated sequencing. The automated sequencing enables the possi- bility of analyzing fetal sequences, at a nucleotide level, in order to detect mutations or polymorphisms which are distinguishable from maternal sequences. KEYWORDS: fetal DNA; maternal blood; mutation; retinitis pigmentosa; automated sequencing INTRODUCTION Circulating cell-free fetal DNA in maternal blood was detected for the first time in 1997 by Lo et al. 1 This discovery opened up a new perspective for the development of noninvasive methods for prenatal diagnosis, thus avoid- ing the risk of fetal loss induced by invasive obstetric procedures. A first aim Address for correspondence: ANA Bustamente, Department of Genetics, Fundacion Jimenez Diaz, Avda. Catolicos 2, 2840 Madrid, Spain. Voice: +34-915504872; fax: +34-915448735. e-mail: abustamente@fjd.es Ann. N.Y. Acad. Sci. 1075: 108–117 (2006). C  2006 New York Academy of Sciences. doi: 10.1196/annals.1368.014 108