The purpose of this Phase II Small Business Innovation Research (SBIR) grant is to develop a new molecular-based method for rapid and low-cost screening for fragile X with broad commercial application. Fragile X syndrome (FXS) is the most common genetic cause of mental retardation in males. FXS is caused by the expansion of DNA sequence via a series of CGG trinucleotide repeats in the FMR1 gene. In normal individuals, the FMR1 gene contains 5 to 45 CGG repeats; however, individuals with FXS have over 200 repeats. Current diagnostic screening for FXS involves molecular assays to determine the size of the repeat segment, but these assays only detect the gross size of CGG repeats and are labor intensive and expensive. In Phase I of this Small Business Innovation Research (SBIR) award, the researchers developed a novel, highly efficient and accurate screening test for diagnosing FXS. In Phase II, the researchers will further optimize this new assay, test it in a clinical diagnostic setting, and test how well the assay works in screening newborn blood spots. This research should lead to the optimization and implementation of a test that is suitable for low cost high-throughput screening for FXS. As such, this test has the potential to markedly change current screening processes for FXS.