Regeneration involves gene expression changes explained in part by context-dependent recruitment of transcriptional activators to distal enhancers. Silencers that engage repressive transcriptional complexes are less studied than enhancers and more technically challenging to validate, but they potentially have profound biological importance for regeneration. Here, we identified candidate silencers through a screening process that examined the ability of DNA sequences to limit injury-induced gene expression in larval zebrafish after fin amputation. A short sequence (s1) on chromosome 5 near several genes that reduce expression during adult fin regeneration could suppress promoter activity in stable transgenic lines and diminish nearby gene expression in knockin lines. High-resolution analysis of chromatin organization identified physical associations of s1 with gene promoters occurring preferentially during fin regeneration, and genomic deletion of s1 elevated the expression of these genes after fin amputation. Our study provides methods to identify “tissue regeneration silencer elements” (TRSEs) with the potential to reduce unnecessary or deleterious gene expression during regeneration. Display omitted •A larval screen identifies candidate silencers during zebrafish fin regeneration•s1 contains sequences essential to repress expression from nearby genes•s1 makes physical contacts with genes that reduce expression during fin regeneration•s1 is essential for regeneration-associated reductions in expression of nearby genes Ando et al. report a screen in zebrafish to identify sequences that repress gene expression during tissue regeneration and find an element with silencing activity that physically contacts nearby genes and tempers their RNA levels during regeneration. The investigation of silencers can illustrate how regeneration programs are controlled in animals.