Warner Yuen, Bobby Concepción, Kate Dungen, Leona Bartell, Toshimasa Watanabe, Kenji Yamamoto
and Eiji Nakamura
Hitachi Software Engineering America Ltd., San Bruno, CA

× Ø × Ø × Ø × Ø × Ø × Ø × Ø × Ø × Ø × Ø × Ø × Ø × Ø × Ø × Ø

Short Tandem Repeat (STR) DNA analysis is ideal for genotyping with instrumentation. STRs can be effectively imaged and analyzed using laser induced fluorescence. One system , the Hitachi FMBIO^®, incorporates a solid state YAG laser for fluorescent excitation and a photo multiplier tube for capture of fluorescent signal. Using this method of detection, we have been able to demonstrate quantitative and qualitative results for STR analysis. The FMBIO^® also has multi-color analysis capabilities that can increase throughput and accuracy in genotyping by using a fluorescent internal size marker in a second color. Ever increasing needs in throughput and discrimination power has led to new developments in software. Software developments include work on new three color analysis software, new genotype scoring software, and high throughput software for CODIS database compatibility.

We will describe the development of new three color analysis software for the FMBIO^®. This new advance allows for greatly enhanced throughput and higher discrimination power in STR systems. In the past, we have used a fluorescent internal size marker in one color and an STR multiplex (CTTv or FFFL) in a second color for increasing throughput. This method greatly improved throughput over traditional silver staining because allelic ladders are no longer required to be run next to every DNA sample, leaving more lanes open for analyzing unknowns.

The new three color software will allow two different color tagged STR sets to be co-migrated with an internal size marker in a third color. The three color software is being tested with new STR systems from both Promega (PowerPlex™) and the British Forensic Science Service (Second Generation Multiplex, SGM). Minimally, this will allow eight STR loci to be amplified and analyzed in a single reaction.

Additionally, we will describe new software, STaR Call, that has been developed for high throughput genotyping and data banking. The genotype scoring software has been designed around a floating bin technique. Consistent sizing and reproducible allele assignments have been demonstrated with this technique from gel to gel and operator to operator. Furthermore, using the FMBIO^® as a loosely coupled CODIS software interface, we have been able to design software compatibility for CODIS. The software has been developed for rapid entry of many sample genotypes at a time into the CODIS PCR/STR database. The architecture has open links that can allow importing of CODIS sample information from an existing database. With further programming, this may also be adapted for bar-code tracking of samples.