Figure 4.

Comparing enhancer activity between human and chimpanzee human accelerated region (HAR) sequences. Massively parallel reporter assay studies involve cloning HAR sequences into reporter vectors along with barcodes that uniquely identify each tested sequence. These vectors are inserted into cell lines, such as neural progenitor cells, using molecular tools such as lentiviruses. They randomly insert into the cell line’s genome. HAR enhancer activity is measured with RNA sequencing of the transcribed barcodes. By associating each tested sequence with many barcodes, activity can be averaged across genomic integration points, providing a robust measurement.