Analysis of fetal DNA from maternal plasma by real-time PCR method offers great potential for noninvasive prenatal genetic diagnosis of sex-linked diseases, rhesus D status, disorders of autosomal dominant inheritance, as well as pregnancy-associated complications. The aim of this study was to evaluate the specificity and sensitivity of real-time quantitative PCR method for fetal gender assessment in early pregnancy (16-20 week of gestation).

Blood samples were collected from 58 pregnant women prior to amniocentesis. DNA was extracted from maternal plasma using a QIAmp DNA Blood Mini Kit. DNA samples were subjected to real-time quantitative PCR amplification of SRY (as a fetus specific marker) and b-globin (as a marker for total plasma DNA) genes.

The b-globin sequence was detected in all samples. SRY gene was detected in 31 of 34 plasma samples from women with male fetuses and in none of the 24 samples from women with female fetuses. Thus, the sensitivity of the real-time quantitative PCR system in our study was 91.2% and the specificity was 100%. The fetal gender was correctly determined in 55 (94.8%) of 58 maternal plasma samples. The concentration of b-globin gene ranged from 161-25.568 GE/ml (median=1136), while the concentration of SRY gene ranged from 5-166 GE/ml (median=63). The percentage of free fetal DNA ranged from 0.1 to 46.1 (median=2.2).

Amplification of free fetal DNA in maternal plasma by real-time quantitative PCR is a promising approach for valid and rapid fetal sex determination in early pregnancy and can be a valuable tool of prenatal diagnosis for pregnant women who are carriers of an X-linked disorder.