A Novel Software Tool for Automated Cluster Calling and Management of TaqMan-Based SNP Genotyping Data

With an ever-increasing number of available genome sequences, single-nucleotide polymorphism (SNP) genotyping is expected soon to become the preferred method for genomics-assisted breeding of most crop plants and farm animals. Currently, TaqMan-based or related technologies using FRET probes (e.g. Molecular Beacons, Invader, Scorpion, etc.) are quickly gaining popularity. These high-throughput methods typically generate massive datasets and therefore require sophisticated analysis and storage solutions. We developed a plugin application for the BioNumerics software, which performs a fully automated cluster calling and provides a flexible user interface for visualization and manual fine-tuning of the calls. The plugin performs its automated cluster calling using a multi-seed partitioning algorithm and reports confidence values per batch and for each individual SNP call. Thresholds for calling, algorithm parameters and report options can be specified by the user. All data, including calls, sample information and experiment settings, are stored in an industry-standard relational database (e.g. SQL Server, Oracle or MySQL). This makes it possible to construct large genotyping databases over long periods of time and allows integration with existing storage platforms or Laboratory Information Management Systems (LIMS). Results are easily visualized in customizable plots and graphs. In addition, SNP scores can be further analyzed in BioNumerics using clustering and dimensioning reduction techniques (PCA, MDS and discriminant analysis) or with statistical tools such as multivariate analysis of variance (MANOVA). In the same software package, SNP genotyping results can be compared and merged with results from other techniques such as microsatellites, AFLP, high-resolution melting curves and phenotypic characterizations.