As submitted by the proposer: Insertions-Deletions (INDELs) are a type of polymorphism where small sequences of DNA have been inserted or deleted in relation to a known consensus reference sequence. The differences between the alleles are based on amplicon size, rather than detecting a nucleotide substitution as used for Single Nucleotide Polymorphism (SNP) typing. These size differences can be easily resolved using capillary electrophoresis (CE) with traditional chemistries for assaying fragment length. Thus, no new instrumentation is required for INDEL analyses in standard forensic laboratories. Analytically, INDELs perform similar to that of STRs and can be multiplexed together to achieve a high power of discrimination, as well as be multiplexed with STRs to facilitate analyses of challenged samples. The amplicon sizes of INDELs can be designed to be short, which are comparable to those of SNPs and are optimal for highly degraded samples. Furthermore, unlike STRs, INDELs do not yield stutter peaks due to slippage during PCR. Thus, interpretation complexity of SNPs can be reduced compare with STRs, especially for Low Copy Number DNA profiles. The mutation rates of INDELs are about 10 times lower than those of the SNPs, which make INDELs desirable for DNA-based kinship analysis. This proposed 2-year project seeks to develop two INDEL panels for human identification and ancestry identification, respectively, based on the data from the 1000 Genomes Project. The 1000 Genomes Project provides the most comprehensive data source for human genome as of today, including more than 1 million short INDELs and ~20,000 long INDELs. The INDELs will be selected with certain population genetics criteria. The INDELs with low allele frequency differences among the populations (measured by Fst) are good for human identification; the INDELs with high Fst are good for ancestry identification. The primers of the INDELs will be designed with a novel primer design approach to have short amplicon sizes (<160bp) to meet forensic application needs. All primers of the markers will be tested in singleplex assays. Furthermore, multiplex assays will be developed based on the selected INDEL panels and will be validated following SWGDAM validation guidelines. The marker selections in both panels will be refined through the developing process. Approximately 40 markers are sought for each panel. The proposed work will provide the forensic community, especially practitioners, new tools to enhance their abilities in analyzing low quantity or highly degraded samples with current standard technology. No new instrumentation is needed to implement these new assays in a standard forensic DNA laboratory. Higher success rates of genotyping can be obtained because of relatively small amplicon sizes and allele length differences based on allele detection. Less complicated interpretation protocols will be needed, since there is no stutter with INDELs. The final product eventually provides an alternative approach for cases with low copy number DNA evidence. In addition, the ancestry identification assay can be used in generating investigative leads and helping solve more cases. This proposed project will be managed by the PI, Dr. Bobby LaRue, who is a Research Assistant Professor in the Institute of Applied Genetics (IAG) at University of North Texas Health Science Center (UNTHSC). Dr. Bruce Budowle, who is the Professor and Executive Director of IAG, and Dr. Jianye Ge, will be co-PI's of this proposed study and contribute throughout the project. Both PI and co-PI's have years of experience in statistical and technical aspects of forensic genetics.