Extended Data Fig. 6. Dissecting mechanisms for how gene loci regulate a neighbor.

(a) Three categories of possible mechanisms by which a gene locus might regulate the expression of a neighbor. (b) We used two strategies to insert pAS downstream of gene promoters. In the first strategy, we inserted a 49-bp synthetic pAS (“spA”) using a single-stranded DNA oligo with 75-bp homology arms (see Methods). (c) In the second pAS insertion strategy, we cloned a donor plasmid containing a selection cassette and three different pAS sequences (see Methods). Homology arms of 300–800 bp were used to integrate the cassette. After isolating clones with successful insertions, we used a second round of transfections to remove the selection cassette, leaving behind three tandem pASs. EFS = elongation factor 1 promoter. Puro = puromycin resistance gene (pac). HSV-tk = herpes simplex virus thymidine kinase.