Objective: Determining fetal sex noninvasively is dependent of a robust assay. We designed a novel SRY assay and combined it with a SRY assay from literature forming a duplex assay with the same fluorescent dye to increase detection of Y-chromosome at low cell-free fetal DNA or chimeric DNA concentrations. Methods: We analyzed 48 plasma samples from pregnant women in gestational weeks 5–35. Fetal sex was determined by DYS14 and SRY. We also compared 2 monoplex SRY assays with a SRY duplex assay at low DNA concentrations. Results: The DYS14 assay was more sensitive than the SRY assay, but less specific at low cell-free fetal DNA concentrations. We obtained a more sensitive duplex SRY assay with the same specificity as the 2 monoplex assays. Conclusion: The SRY duplex assay improves the SRY detection at low DNA concentration.

Keywords:
Y-chromosome, SRY, Fetal marker, Prenatal, Real-time PCR, NIPD
1.
Lo YM, Corbetta N, Chamberlain PF, Rai V, Sargent IL, Redman CW, Wainscoat JS: Presence of fetal DNA in maternal plasma and serum. Lancet 1997;350:485–487.
2.
Lo YM, Tein MS, Lau TK, Haines CJ, Leung TN, Poon PM, Wainscoat JS, Johnson PJ, Chang AM, Hjelm NM: Quantitative analysis of fetal DNA in maternal plasma and serum: implications for noninvasive prenatal diagnosis. Am J Hum Genet 1998;62:768–775.
3.
Illanes S, Denbow M, Kailasam C, Finning K, Soothill PW: Early detection of cell-free fetal DNA in maternal plasma. Early Hum Dev 2007;83:563–566.
4.
Benachi A, Steffann J, Gautier E, Ernault P, Olivi M, Dumez Y, Costa JM: Fetal DNA in maternal serum: does it persist after pregnancy? Hum Genet 2003;113:76–79.
5.
Alberry M, Maddocks D, Jones M, Abdel HM, Abdel-Fattah S, Avent N, Soothill PW: Free fetal DNA in maternal plasma in anembryonic pregnancies: confirmation that the origin is the trophoblast. Prenat Diagn 2007;27:415–418.
6.
Flori E, Doray B, Gautier E, Kohler M, Ernault P, Flori J, Costa JM: Circulating cell-free fetal DNA in maternal serum appears to originate from cyto- and syncytio-trophoblastic cells. Case report. Hum Reprod 2004;19:723–724.
7.
Masuzaki H, Miura K, Yoshiura KI, Yoshimura S, Niikawa N, Ishimaru T: Detection of cell free placental DNA in maternal plasma: direct evidence from three cases of confined placental mosaicism. J Med Genet 2004;41:289–292.
8.
Tjoa ML, Cindrova-Davies T, Spasic-Boskovic O, Bianchi DW, Burton GJ: Trophoblastic oxidative stress and the release of cell-free feto-placental DNA. Am J Pathol 2006;169:400–404.
9.
Chan KC, Zhang J, Hui AB, Wong N, Lau TK, Leung TN, Lo KW, Huang DW, Lo YM: Size distributions of maternal and fetal DNA in maternal plasma. Clin Chem 2004;50:88–92.
10.
Li Y, Zimmermann B, Rusterholz C, Kang A, Holzgreve W, Hahn S: Size separation of circulatory DNA in maternal plasma permits ready detection of fetal DNA polymorphisms. Clin Chem 2004;50:1002–1011.
11.
Fan HC, Blumenfeld YJ, Chitkara U, Hudgins L, Quake SR: Noninvasive diagnosis of fetal aneuploidy by shotgun sequencing DNA from maternal blood. Proc Natl Acad Sci USA 2008;105:16266–16271.
12.
Scheffer PG, van der Schoot CE, Page-Christiaens GC, Bossers B, van Erp F, de Haas M: Reliability of fetal sex determination using maternal plasma. Obstet Gynecol 2010;115:117–126.
13.
Rijnders RJ, Van Der Luijt RB, Peters ED, Goeree JK, Van Der Schoot CE, Ploos Van Amstel JK, Christiaens GC: Earliest gestational age for fetal sexing in cell-free maternal plasma. Prenat Diagn 2003;23:1042–1044.
14.
Wright CF, Burton H: The use of cell-free fetal nucleic acids in maternal blood for non-invasive prenatal diagnosis. Hum Reprod Update 2009;15:139–151.
15.
Hyland CA, Gardener GJ, Davies H, Ahvenainen M, Flower RL, Irwin D, Morris JM, Ward CM, Hyett JA: Evaluation of non-invasive prenatal RHD genotyping of the fetus. Med J Aust 2009;191:21–25.
16.
Finning KM, Martin PG, Soothill PW, Avent ND: Prediction of fetal D status from maternal plasma: introduction of a new noninvasive fetal RHD genotyping service. Transfusion 2002;42:1079–1085.
17.
Wang XD, Wang BL, Ye SL, Liao YQ, Wang LF, He ZM: Non-invasive foetal RHD genotyping via real-time PCR of foetal DNA from Chinese RhD-negative maternal plasma. Eur J Clin Invest 2009;39:607–617.
18.
Ekelund CK, Skibsted L, Sogaard K, Main KM, Dziegiel MH, Schwartz M, Moeller N, Roos L, Tabor A: Dizygotic monochorionic twin pregnancy conceived following intracytoplasmic sperm injection treatment and complicated by twin-twin transfusion syndrome and blood chimerism. Ultrasound Obstet Gynecol 2008;32:832–834.
19.
Picchiassi E, Coata G, Fanetti A, Centra M, Pennacchi L, Di Renzo GC: The best approach for early prediction of fetal gender by using free fetal DNA from maternal plasma. Prenat Diagn 2008;28:525–530.
20.
Finning KM, Chitty LS: Non-invasive fetal sex determination: impact on clinical practice. Semin Fetal Neonatal Med 2008;13:69–75.
21.
Avent ND, Chitty LS: Non-invasive diagnosis of fetal sex; utilisation of free fetal DNA in maternal plasma and ultrasound. Prenat Diagn 2006;26:598–603.
22.
Zimmermann BG, Holzgreve W, Avent N, Hahn S: Optimized real-time quantitative PCR measurement of male fetal DNA in maternal plasma. Ann N Y Acad Sci 2006;1075:347–349.
23.
Zhong XY, Holzgreve W, Hahn S: Direct quantification of fetal cells in maternal blood by real-time PCR. Prenat Diagn 2006;26:850–854.
24.
Hill M, Finning K, Martin P, Hogg J, Meaney C, Norbury G, Daniels G, Chitty LS: Non-invasive prenatal determination of fetal sex: translating research into clinical practice. Clin Genet 2011;80:68–75.
25.
Zimmermann B, El-Sheikhah A, Nicolaides K, Holzgreve W, Hahn S: Optimized real-time quantitative PCR measurement of male fetal DNA in maternal plasma. Clin Chem 2005;51:1598–1604.
26.
Clausen FB, Krog GR, Rieneck K, Rasmark EE, Dziegiel MH: Evaluation of two real-time multiplex PCR screening assays detecting fetal RHD in plasma from RhD negative women to ascertain the requirement for antenatal RhD prophylaxis. Fetal Diagn Ther 2011;29:155–163.
27.
Petersen B, Lundegaard C, Petersen TN: NetTurnP – neural network prediction of beta-turns by use of evolutionary information and predicted protein sequence features. PLoS One 2010;5:e15079.
28.
Horinek A, Korabecna M, Panczak A, Ulcova GZ, Nouzova K, Calda P, Hancarova M: Cell-free fetal DNA in maternal plasma during physiological single male pregnancies: methodology issues and kinetics. Fetal Diagn Ther 2008;24:15–21.
29.
Hromadnikova I, Houbova B, Hridelova D, Voslarova S, Kofer J, Komrska V, Habart D: Replicate real-time PCR testing of DNA in maternal plasma increases the sensitivity of non-invasive fetal sex determination. Prenat Diagn 2003;23:235–238.
30.
Johnson KL, Dukes KA, Vidaver J, Leshane ES, Ramirez I, Weber WD, Bischoff FZ, Hahn S, Sharma A, Dang DX, Hire LM, Bianchi DW, Simpson JL, Holzgreve W, Elias S, Klinger KW: Interlaboratory comparison of fetal male DNA detection from common maternal plasma samples by real-time PCR. Clin Chem 2004;50:516–521.
31.
Segal E, Fondufe-Mittendorf Y, Chen L, Thastrom A, Field Y, Moore IK, Wang JP, Widom J: A genomic code for nucleosome positioning. Nature 2006;442:772–778.
32.
Zhang Z, Pugh BF: High-resolution genome-wide mapping of the primary structure of chromatin. Cell 2011;144:175–186.
© 2012 S. Karger AG, Basel
2012
Copyright / Drug Dosage / Disclaimer
Copyright: All rights reserved. No part of this publication may be translated into other languages, reproduced or utilized in any form or by any means, electronic or mechanical, including photocopying, recording, microcopying, or by any information storage and retrieval system, without permission in writing from the publisher.
Drug Dosage: The authors and the publisher have exerted every effort to ensure that drug selection and dosage set forth in this text are in accord with current recommendations and practice at the time of publication. However, in view of ongoing research, changes in government regulations, and the constant flow of information relating to drug therapy and drug reactions, the reader is urged to check the package insert for each drug for any changes in indications and dosage and for added warnings and precautions. This is particularly important when the recommended agent is a new and/or infrequently employed drug.
Disclaimer: The statements, opinions and data contained in this publication are solely those of the individual authors and contributors and not of the publishers and the editor(s). The appearance of advertisements or/and product references in the publication is not a warranty, endorsement, or approval of the products or services advertised or of their effectiveness, quality or safety. The publisher and the editor(s) disclaim responsibility for any injury to persons or property resulting from any ideas, methods, instructions or products referred to in the content or advertisements.
You do not currently have access to this content.