DNA oligonucleotides are essential components of a high number of technologies in molecular biology. The key event of each oligonucleotide-based assay is the specific binding between oligonucleotides and their target DNA. However, single-stranded DNA molecules also tend to bind to unintended targets or themselves. The probability of such unspecific binding increases with the complexity of an assay. Therefore, accurate data management and design workflows are necessary to optimize the in-silico design of primers and probes. Important considerations concerning computational infrastructure and run time need to be made for both data management and the design process. Data retrieval, data updates, storage, filtering and analysis are the main parts of a sequence data management system. Each part needs to be well-implemented as the resulting sequences form the basis for the oligonucleotide design. Important key features, such as the oligonucleotide length, melting temperature, secondary structures and primer dimer formation, as well as the specificity, should be considered for the in-silico selection of oligonucleotides. The development of an efficient oligonucleotide design workflow demands the right balance between the precision of the applied computer models, the general expenditure of time, and computational workload. This paper gives an overview of important parameters during the design process, starting from the data retrieval, up to the design parameters for optimized oligonucleotide design.
Keywords: Alignment; Oligonucleotide design; Oligonucleotide secondary structures; Sequence database.