High-Throughput Single Nucleotide Mutagenesis

Many research groups are interested in developing high-throughput methods for dissecting the functional impact of genetic variation in their genes or non-coding regions of interest. The production of libraries of DNA molecules, each different from the reference by one and only one position currently requires the purchase of many thousands of synthesized oligonucleotides either to be directly cloned into a vector, used as the donor DNA with CRISPR-Cas9 or used as primers in multiplex mutagenesis protocol . We have developed a method of massively parallel single nucleotide mutagenesis in which such libraries can be made in a single day for<$30/RXN. The method was recently accepted for publication at Nature Methods! We are currently using this method to functionally assess ALL POSSIBLE single nucleotide changes in several Mendelian disease genes to predict which variants are capable of causing disease!