Since the literature contains references to Y-STR multiplexes that at most amplify only five or six loci simultaneously, the authors of this article have attempted to increase the level of multiplexing for Y STRs.
This effort has successfully demonstrated a Y-STR 10-plex, thus providing a megaplex capable of simultaneously amplifying 20 different Y-STR markers. The megaplex includes all of the Y-STRs that compose the "extended haplotype" used in Europe (DYS19, DYS385; DYS389/II, DYS 3891/II, DYS390, DYS391, DYS392, DYS393, and YCAII). Plus additional polymorphic Y-STRs DYS437, DYS438, DYS439, DYS447, DYYS448, DYS388, DYS426, GATA A7.1, and GATA H4. The authors are also developing SNP assays from the Y-chromosome that may be rapidly analyzed with time-of-flight mass spectrometry. With improved multiplexes and techniques for more rapid data collection, Y-chromosome haplotypes may be more quickly generated for DNA databases, and forensic cases may be solved more efficiently. In order to develop new multiplex Y-STR assays, schematic layouts were made to illustrate potential allele size ranges and dye labels for each marker. The allele size ranges were defined through extensive literature searches and testing of the Y-chromosome consortium panel. Primers were than designed using Primer 3 with defined PCR product sizes to match the required allele range. Candidate primers were screened for potential primer cross-reactions using a custom-designed program written in Visual Basic. Samples were run on both the ABI 310 and 3100 Genetic Analyzers, using manufacturer recommended electrophoresis conditions and appropriate matrix standards. Multiplex PCR was performed with 5 Y-SNP markers. Primer extension reactions were then performed using a Bruker BIFLEX III time-of-flight mass spectrometer in reflectron mode. 3 figures and 13 references