Sample stacking capillary electrophoretic microdevice for highly sensitive mini Y short tandem repeat genotyping

Abstract

Lab-on-a-chip provides an ideal platform for short tandem repeat (STR) genotyping due to its intrinsic low sample consumption, rapid analysis, and high-throughput capability. One of the challenges, however, in the forensic human identification on the microdevice is the detection sensitivity derived from the nanoliter volume sample handling. To overcome such a sensitivity issue, here we developed a sample stacking CE microdevice for mini Y STR genotyping. The mini Y STR includes redesigned primer sequences to generate smaller-sized PCR amplicons to enhance the PCR efficiency and the success rate for a low copy number and degraded DNA. The mini Y STR amplicons occupied in the 5- and 10-mm stacking microchannels are preconcentrated efficiently in a defined narrow region through the optimized sample stacking CE scheme, resulting in more than tenfold improved fluorescence peak intensities compared with that of a conventional cross-injection microcapillary electrophoresis method. Such signal enhancement allows us to successfully analyze the Y STR typing with only 25 pg of male genomic DNA, with high background of female genomic DNA, and with highly degraded male genomic DNA. The combination of the mini Y STR system with the novel sample stacking CE microdevice provides the highly sensitive Y STR typing on a chip, making it promising to perform high-performance on-site forensic human identification.