The application of array-based oligonucleotides in biological studies is described. These oligonucleotides are mainly used to design large libraries of various nucleotide sequences, which are applied to study protein-nucleic acid interactions, splicing, transcription, translation, and other regulatory processes in mammalian, yeast, and bacterial systems. The application of gene libraries generated by array-based nucleotides along with advanced methods of the combination of DNA duplexes will make it possible to obtain complex genetic designs for synthetic biology.