Sex Determination by PCR Analysis of the X-Y Amelogenin Gene

Abstract

PCR-based sex determination was first accomplished by amplifying multiple-copy sequences in the Y-chromosomal DYZ1 locus (1,2). These methods are quite sensitive (amplifiable from trace samples) due to the multiplicity, but it is impossible to tell whether the template DNA is from a female or whether the analysis has ended in failure when no fragment is amplified by PCR. Simultaneous amplification of X- and Y-chromosomal genes is thus necessary for a reliable sex test, in which the X-specific product may act as an internal control for PCR. From this point of view, centromeric alphoid repeats on the X and Y chromosomes have been amplified separately (3) or in the same reaction mixture (4) using two pairs of primers. These methods are also sensitive. However, a difference in the copy numbers of the repeated sequences (i.e., 5000 and 100 copies in X and Y centromeres, respectively) is likely to cause a difference in the sensitivity of PCR amplification so that the X product does not act strictly as an internal control. Sex determination by PCR analysis of X-Y homologous genes is thus most trustworthy, because X and Y sequences are of equal (single) copy number and because they can be amplified using one set of primers.