Functional assessment of disease-associated regulatory variants in vivo using a versatile dual colour transgenesis strategy in zebrafish.
PLoS Genet
; 11(6): e1005193, 2015 Jun.
Article
en En
| MEDLINE
| ID: mdl-26030420
ABSTRACT
Disruption of gene regulation by sequence variation in non-coding regions of the genome is now recognised as a significant cause of human disease and disease susceptibility. Sequence variants in cis-regulatory elements (CREs), the primary determinants of spatio-temporal gene regulation, can alter transcription factor binding sites. While technological advances have led to easy identification of disease-associated CRE variants, robust methods for discerning functional CRE variants from background variation are lacking. Here we describe an efficient dual-colour reporter transgenesis approach in zebrafish, simultaneously allowing detailed in vivo comparison of spatio-temporal differences in regulatory activity between putative CRE variants and assessment of altered transcription factor binding potential of the variant. We validate the method on known disease-associated elements regulating SHH, PAX6 and IRF6 and subsequently characterise novel, ultra-long-range SOX9 enhancers implicated in the craniofacial abnormality Pierre Robin Sequence. The method provides a highly cost-effective, fast and robust approach for simultaneously unravelling in a single assay whether, where and when in embryonic development a disease-associated CRE-variant is affecting its regulatory function.
Texto completo:
1
Colección:
01-internacional
Banco de datos:
MEDLINE
Asunto principal:
Síndrome de Pierre Robin
/
Transgenes
/
Elementos Reguladores de la Transcripción
Tipo de estudio:
Prognostic_studies
/
Risk_factors_studies
Límite:
Animals
Idioma:
En
Revista:
PLoS Genet
Asunto de la revista:
GENETICA
Año:
2015
Tipo del documento:
Article
País de afiliación:
Reino Unido