Real-Time PCR-Based Screening for Homozygous SMN2 Deletion Using Residual Dried Blood Spots

The survival motor neuron 2 ( ) gene is a recognized modifier gene of spinal muscular atrophy (SMA). However, our knowledge about the role of -other than its modification of SMA phenotypes-is very limited. Discussions regarding the relationship between homozygous deletion and motor neuron diseases, including amyotrophic lateral sclerosis, have been mainly based on retrospective epidemiological studies of the diseases, and the precise relationship remains inconclusive. In the present study, we first estimated that the frequency of homozygous deletion was ~1 in 20 in Japan. We then established a real-time polymerase chain reaction (PCR)-based screening method using residual dried blood spots to identify infants with homozygous deletion. This method can be applied to a future prospective cohort study to clarify the relationship between homozygous deletion and motor neuron diseases. In our real-time PCR experiment, both PCR (low annealing temperatures) and blood (high hematocrit values and low white blood cell counts) conditions were associated with incorrect results (i.e., false negatives and positives). Together, our findings not only help to elucidate the role of , but also aid in our understanding of the pitfalls of current SMA newborn screening programs for detecting homozygous deletions.