Improving DNA data exchange: validation studies on a single 6 dye STR kit with 24 loci.

The idea of developing a new multiplex STR amplification system was conceived in 2011 as an effective way to implement the new European standard set (ESS) of 12 STR markers adopted by The Council of the European Union in 2009 while maintaining an effective compatibility and information exchange with the historical DNA profiles contained in the Spanish national DNA database (around 200,000 DNA profiles) mainly based on the 13 CODIS core STR loci plus D19S433 and D2S1338 markers. With this goal in mind we proposed to test and validate a single STR amplification system for simultaneous analysis of 21 STR markers covering both CODIS and ESS core STR loci plus three additional markers (D19S433, D2S1338, and SE33) also contained in commonly used STR kits and national DNA databases. In 2012, we started the first beta-testing with a 6-dye STR kit prototype containing 24 loci (now known as the GlobalFiler™ PCR Amplification Kit) developed by Life Technologies in response to the CODIS Core Loci Working Group's recommendation to expand the CODIS Core Loci.

Challenges of DNA profiling in mass disaster investigations.

In cases of mass disaster, there is often a need for managing, analyzing, and comparing large numbers of biological samples and DNA profiles. This requires the use of laboratory information management systems for large-scale sample logging and tracking, coupled with bioinformatic tools for DNA database searching according to different matching algorithms, and for the evaluation of the significance of each match by likelihood ratio calculations. There are many different interrelated factors and circumstances involved in each specific mass disaster scenario that may challenge the final DNA identification goal, such as: the number of victims, the mechanisms of body destruction, the extent of body fragmentation, the rate of DNA degradation, the body accessibility for sample collection, or the type of DNA reference samples availability.

Real-time PCR designs to estimate nuclear and mitochondrial DNA copy number in forensic and ancient DNA studies.

We explore different designs to estimate both nuclear and mitochondrial human DNA (mtDNA) content based on the detection of the 5' nuclease activity of the Taq DNA polymerase using fluorogenic probes and a real-time quantitative PCR detection system. Human mtDNA quantification was accomplished by monitoring the real-time progress of the PCR-amplification of two different fragment sizes (113 and 287 bp) within the hypervariable region I (HV1) of the mtDNA control region, using two fluorogenic probes to specifically determine the mtDNA copy of each fragment size category. This mtDNA real-time PCR design has been used to assess the mtDNA preservation (copy number and degradation state) of DNA samples retrieved from 500 to 1500 years old human remains that showed low copy number and highly degraded mtDNA.

Specific quantification of human genomes from low copy number DNA samples in forensic and ancient DNA studies.

We reviewed the current methodologies used for human DNA quantitation in forensic and ancient DNA studies, including sensitive hybridization methods based on the detection of nuclear alpha-satellite repetitive DNA regions or more recently developed fluorogenic real-time polymerase chain reaction (PCR) designs for the detection of both nuclear and mitochondrial DNA regions. Special emphasis has been put on the applicability of recently described different real-time PCR designs targeting different fragments of the HV1 mtDNA control region, and a segment of the X-Y homologous amelogenin gene. The importance of these quantitative assays is to ensure the consistency of low copy number DNA typing (STR profiling and mtDNA sequencing).