Genotype validation of reciprocal genomic disorder CRISPR lines

Recurrent genomic disorders (RGDs) occur in ~1 in 200 newborns. These syndromes are caused by deletions and duplications in the DNA that are created when repetitive low copy repeats of a genomic segment misalign via Non-Allelic Homologous Recombination (NAHR). Some phenotypic characteristics are specific to the rearranged locus, for example, hyperphagia in 15q11.2-13 paternal deletion or overgrowth syndrome in 5q35 deletion. However, RGDs in general are among the most common recurrent cases of neurodevelopmental disorders, although originating from different genomic loci. The reason for these shared pathogenic processes remains unknown.

Using CRISPR techniques we targeted low copy repeats in 8p23 and 7q11 to generate deletions and duplications in induced pluripotent stem cells (iPSC). The goal of my research stay was to validate the RGD genotypes of the iPSC via copy number determination using droplet digital PCR. Afterwards, we differentiated the isogenic RGD lines to induced neurons to study their neurodevelopmental characteristics via RNA sequencing and electrophysiological assays. Identifying connections in the genetic consequences of RGDs will shed light on why they can often originate from different mutations but cause similar phenotypic characteristics.

For more information: hmwilliams@eckerd.edu