The authors want to identify the major molecules involved in synaptogenesis. They discovered sushi repeat-containing protein X-linked 2 (SRPX2) which modulates the synapse density in hippocampal neurons. Mutations in this gene were identified in patients suffering of epilepsy with associated oral and speech dyspraxia. In the search for synaptogenic factors, they performed a high-throughput screen of genes encoding for membrane and secreted proteins that mediate synaptogenesis in the CNS. Neuronal cultures were transfected with individual genes using the Nucleofector™ 96-well Shuttle™ System, and then assessed for excitatory and inhibitory synaptic density in transfected neurons. They identified SRPX2 as a modulator of excitatory synapse density. Neurons cells coming from dissociated hippocampus were transfected in some of the experiments with liposome-based reagents but low transfection efficiency was obtained. Highly efficient transfection of both shRNA and DNA plasmids was obtained when Nucleofector™ 96-well Shuttle™ System was used. By a series of overexpression or inhibition experiments of SRPX2 levels, they manage to demonstrate the role of this protein in the excitatory synapses formation. Moreover, the authors showed that in hippocampus, SRPX2 levels are strictly regulated by Fox2 transcription factor and a dysregulation of SPRX2 levels results in impaired ultrasonic vocalization in infant mice.