Robustness of the YSTRs DYS19, DYS389 I and II, DYS390 and DYS393: optimization of a PCR pentaplex

Various technical methods were investigated with the aim of developing a mu ltiplex system to amplify five Y-chromosome STR loci in the same PCR reacti on: DYS393, DYS19, DYS390, DYS389 I and DYS389 II. A sequenced allelic ladd er was constructed with previously sequenced alleles including the most com mon ones. A number of reamplification conditions of the allelic ladders wer e tested. The pentaplex was evaluated for typing using two different platfo rms (ABI and ALF) with promising results. However, in degraded samples non- specific artifacts were observed in the DYS393 system in the same range of sizes as the real alleles. This system can also be typed in females under r elatively low stringency conditions in the PCR amplification, making this s ystem prone to errors in critical samples. This lack of specificity can be reduced by increasing the stringency of the PCR conditions. The DYS19 ladde r cannot be reamplified as stutters appear after a few reamplifications. Th ese stutters are probably due to a 2 bp slippage induced by the presence of a TA repeat stretch in the PCR amplified fragments. Non-specific products were also noted in the DYS389 I and DYS389 II amplification, although out o f the range of other alleles in this pentaplex. This newly constructed pent aplex has proved to be very useful in population genetic studies because al l five Y STR markers can be loaded in the same lane of a gel with other Y S TR singleplex or multiplexes. The usefulness of Y-chromosome STRs in crimin al casework is especially evident in analyzing azoospermic individuals.(C) 1999 Elsevier Science Ireland Ltd. All rights reserved.